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THE INTERNATIONAL JOURNAL OF FUNGAL TAXONOMY & NOMENCLATURE

VOLUME 130(1) JANUARY-MARCH 2015

Lactarius olivaceoglutinus sp. nov.
(Das, Verbeken & Nuytinck— PLareE 4, p. #115)
Kanab Das, artist

ISSN (PRINT) 0093-4666 http://dx.doi.org/10.5248/130 | ISSN (ONLINE) 2154-8889
MYXNAE 130(1): 1-306 (2015)

EDITORIAL ADVISORY BOARD

SABINE HUHNDORE (2011-2016) , Chair
Chicago, Illinois, U.S.A.
Scott A. REDHEAD (2010-2015), Past Chair
Ottawa, Ontario, Canada
PETER BUCHANAN (2011-2017)
Auckland, New Zealand

BRANDON MATHENY (2013-2018)
Knoxville, Tennessee, U.S.A.

KAREN HANSEN (2014-2019)
Stockholm, Sweden

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© Mycotaxon, LTp, 2015

MY COTA XON

THE INTERNATIONAL JOURNAL OF FUNGAL TAXONOMY & NOMENCLATURE

VOLUME 130(1)

JANUARY-MARCH 2015

EDITOR-IN-CHIEF

LORELEI L. NORVELL

editor@mycotaxon.com
Pacific Northwest Mycology Service
6720 NW Skyline Boulevard
Portland, Oregon 97229-1309 USA

NOMENCLATURE EDITOR

SHAUN R. PENNYCOOK

PennycookS@LandcareResearch.co.nz
Manaaki Whenua Landcare Research
Auckland, New Zealand

CONSISTING OF I-X + 306 PAGES INCLUDING FIGURES

ISSN 0093-4666 (PRINT) http://dx.doi.org/10.5248/130(1).cvr ISSN 2154-8889 (ONLINE)

Iv ... MYCOTAXON 130(1)

MY COTAXON

VOLUME ONE HUNDRED THIRTY (1) — TABLE OF CONTENTS

COVER SECTION

EPA inch on okt il sna eRe Meebo oy Aicch OORT AL Sey Dente cA Mea ina At eae RR apna vale 4 vi
ROVICWICTS ENE ry oh OE eee cE EEE Rh Eth Ae hee ee EAS Oke ete es vii
SOI TSSIOMDTOCCHUMES 3 25 vi Peurizh date ths Red ng er a Ma meri decease 8 Ie Viii
ERO TE PEEL Mosty BF Soy inn Pie de On ay wan, A hey nme MERE ix

RESEARCH ARTICLES

Pseudoplectania lignicola sp. nov. described from central Europe
S. GLEJDURA, V. KUCERA, P. LIZON, & V. KUNCA
Additions to rust and chytrid pathogens of Turkey
CuMALI OzasLAN, MAKBULE ERDOGDU, ELsAD HUSEYIN, & ZEKIYE SULUDER
Dentipellicula austroafricana sp. nov. supported by morphological
and phylogenetic analyses —‘Jta-JIA CHEN, Lu-Lu SHEN, & YU-CHENG Dal
A new species of Terriera (Rhytismatales, Ascomycota) on
Photinia villosa Quine Li, YuaN Wu, Dan-Dan Lu, YA-Fer Xu, & YING-REN LIN
New records of one Amygdalaria and three Porpidia species
(Lecideaceae) from China Lu-Lu ZHANG, XIN ZHAO & Line Hu

Anungitea guangxiensis and Ellisembia longchiensis, two new species
from southern China JI-WEN X1A, YING-Ru1 Ma, & X1u-GUo ZHANG
Fistulina subhepatica sp. nov. from China inferred from
morphological and sequence analyses
JE SonG, MEI-LING HAN, & BAo-Kat CuI
Antherospora sukhomlyniae, a new species of smut fungi on
Hyacinthella in Crimea (Ukraine) KyryLo G. SAVCHENKO
Tuber xanthomonosporum, a new Paradoxa-like species from China
YUAN QING, SHU-HONG LI, CHENG-YI Liu, LIN Li, MEI YANG,
XIAO-LEI ZHANG, XIAO-LIN LI, LIN-YONG ZHENG, & YUN WANG
Distance1D - a protein profile analytical program designed for
fungal taxonomy
DuSAN MATERIC, BILJANA KUKAVICA, & JELENA VUKOJEVIC
Coccomyces prominens sp. nov. (Rhytismataceae) on Rhododendron
coeloneurum in China Ya-FeI Xu, YUAN Wu, YAN-QIONG MENG,
SHI-JUAN WANG, & YING-REN LIN

Type studies on Amauroderma species described by J.D. Zhao et al.
and the phylogeny of species in China MENG-Jrz Li & Hal-SHENG YUAN

JANUARY-MARCH 2015... V

Cladonia corymbescens consists of two species
TEUVO AHTI, RAQUEL PINO-Bopas, & SOILISTENROOS 91
Morphology and phylogeny of four new Lactarius species
from Himalayan India
KANAD Das, ANNEMIEKE VERBEKEN, & JORINDE NUYTINCK 105

Synonymy of two species of Bipolaris from aquatic crops of Poaceae
ZI-LAN XIAO, KEVIN D. HYDE, & JING-ZE ZHANG 131
Myxomycetes of Sonora (Mexico) 6. Central plains of the Sonoran Desert
Marcos LIZARRAGA, GABRIEL MORENO, MARTIN ESQUEDA,
CYNTHIA SALAZAR-MARQUEZ, & MARTHA L. CORONADO 145

New taxa of Ambomucor (Mucorales, Mucoromycotina)
from China X1A0-YONG Liu & RU-YONG ZHENG 165

Perenniporia koreana, a new wood-rotting basidiomycete
from South Korea YEONGSEON JANG, SEOKYOON JANG,
YOUNG WOON LIM, CHANGMU KIM, & JAE-JIN Kim 173
New records of corticolous myxomycetes from Turkey
R. BATUR ORAN & C. CEM ErGut 181
A new species of Lophodermium with variously branched paraphyses
Hat-Lin Gu, YA-FEI Xu, DAN-DAN LU, SHI-JUAN WANG, & YING-REN LIN 191
Bertia hainanensis sp. nov. (Coronophorales) from southern China
LARISSA N. VASILYEVA, HaI-X1A Ma,
ALEKSEY V. CHERNYSHEV, & STEVEN L. STEPHENSON 197
Distribution of Alternaria species among sections. 1.
Section Porri PHILIpp B. GANNIBAL 207
Morchella galilaea, an autumn morel from Turkey
HATIRA TASKIN, HASAN HUtsryvIn DoGAN, & SAADET BUYUKALACA 215
Hyphoderma hallenbergii, a new corticioid species from India
MANINDER Kaur, AVNEET P. SINGH, & G.S. DHINGRA 223
Xylaria thailandica- a new species from southern Thailand
NATTHAPACH SRIHANANT, VASUN PETCHARAT, & LARISSA N. VASILYEVA 227
Rosellinia brunneola sp. nov. and R. beccariana new to China
WEI LI & Lin Guo 233
Phyllachora hainanensis sp. nov. from China
Na Liu, LE Wana, Gu Huang, & LIN Guo 237
Two new species of Zasmidium from Nepal RAVINDRA NATH KHARWAR,
ARCHANA SINGH, RAGHVENDRA SINGH, & SHAMBHU KuMAR 241
The lichen genus Leiorreuma in China
X1A0-Hua WANG, LI-LI XU, & ZE-FENG JIA 247

VI ... MYCOTAXON 130(1)

Periconiella liquidambaricola sp. nov. - a new Chinese hyphomycete
Uwe BrAuN, STEFFEN BIEN, Lyp1a HONIG, & BETTINA HEUCHERT 253
First record of Erysiphe syringae-japonicae in Turkey
ILGaz AKATA & VASYL P. HELUTA 259
AFLP characterization of three argentine Coprotus species
ARACELI MARCELA RAMOs, LuIS FRANCO TADIC,
NAHUEL POLICELLI, LAURA INES FERREYRA, & ISABEL ESTHER CINTO 265
Verticicladus hainanensis, a new aquatic hyphomycete
MiNG-TIAN Guo, MIN QIAO, JIAN- YING Li, WEI WANG, & ZE-FEN YU 275
Geographic distribution of Sarcoporia polyspora and
Sarcoporia longitubulata sp. nov.
JOSEF VLASAK, JOSEF VLASAK JR., JUHA KINNUNEN, & VIACHESLAV SPIRIN 279
Coccomyces neolitseae sp. nov. from Zhangjiajie, China
QinG Li, YU-X1A CHEN, CHUN-TAO ZHENG, DAN-DAN LU, & YING-REN LIN 289
Datronia ustulatiligna sp. nov. (Agaricomycetes) from India
HARPREET KAUR, GURPREET Kaur, & G.S. DHINGRA 295
Three new species of Xylaria from China
Gu Huana, RuisHa WANG, LIN Guo, & Na Liu 299
NOMENCLATURAL NOVELTIES AND TYPIFICATIONS
PROPOSED IN MYCOTAXON 130-1 305

ERRATA FROM PREVIOUS VOLUMES

VOLUME 129

p-293, line 2 FOR: HASAN HUSEYIN DOGAN READ: HASAN HUSEYIN DOGAN

p.296, line 32 FOR: Lado, C. READ: Lado C.

p.455, line 28 FOR: DM100 READ: DM1000

p.457, line 3 FOR: Spores ornamented with long ridges, arranged in an incomplete banded
reticulum.

READ: Spores were decorated with long ridges sometimes forming an
incomplete banded reticulum.
p.457, line 17 FOR: (ca. 9-12 um diam.) and unbranched and thicker
READ: (ca. 9-12 um diam.) and thicker
p.458, line 9 FOR: National Basic Research Program of China (3140010180)
READ: National Basic Research Program of China (3140010180) and National
Science and Technology Foundation Project (2014FY210400)

JANUARY-MARCH 2015 ...

REVIEWERS — VOLUME ONE HUNDRED THIRTY (1)

The Editors express their appreciation to the following individuals who have,
prior to acceptance for publication, reviewed one or more of the papers

prepared for this volume.

André Aptroot

N.S. Atri

Z.M. Azbukina
Gerald L. Benny

Lina Bettucci

Uwe Braun

Ana Rosa Burgaz

Lei Cai

Matteo Carbone

Lori Carris

Rafael FE Castaneda- Ruiz
Julia Checa

Gilberto Coelho
Roger T.A. Cook
Bao-Kai Cui
Yu-Cheng Dai
Alejandra Teresa Fazio
Ricardo Galan

Tatiana Baptista Gibertoni
Shouyu Guo

Ian Robert Hall

Nils Hallenberg
Tsutomu Hattori
Vasyl Heluta

Bo Huang

Santosh Joshi

M. Kakishima
Mitko Karadelev
Bryce Kendrick
Daniel P. Lawrence
Hua-jie Liu

Xiao- Yong Liu

Dimuthu S. Manamgoda

Milan Matavulj

Eric H.C. McKenzie
V.A. Melnik

David W. Mitchell
Wieslaw Mulenko
Lorelei L. Norvell
Aysun Peksen
Shaun R. Pennycook

Liliane Petrini

Marcin Piatek
Michele D. Piercey-
Normore

Chaninun Pornsuriya
Adam Rollins

Daniel J. Royse

B.M. Sharma

Roger G. Shivas
Mirjana Stajic

Steven L. Stephenson
Guangyu Sun

Joanne E. Taylor
Michal TomSovsky
Yuri Tykhonenko
Larissa Vasilyeva
Xiang-Hua Wang
Zheng Wang

A.J.S. Whalley

Ming Ye

Alessandra Zambonelli

Xiu-Guo Zhang
Li-Wei Zhou

PUBLICATION DATE FOR VOLUME ONE HUNDRED TWENTY-NINE (2)

MYCOTAXON for OcTOBER-DECEMBER 2014, (I-VI + 215-495)

was issued on January 8, 2015

VII

vill ... MYCOTAXON 130(1)

FOUR STEPS TO SUCCESSFUL MYCOTAXON PUBLICATION IN 2015

Prospective MycotTaxon authors should download instructions PDF, review and
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MYCOTAXON ONLINE— www.ingentaconnect.com/content/mtax/mt
Mycotaxon publishes four quarterly issues per year. Both open access and
subscription articles are offered.

JANUARY-MARCH 2015 ... IX

FROM THE EDITOR-IN-CHIEF

WELCOME TO THE YEAR OF MyYCOTAXON 130! — For several decades we have
determined MycoTaxon volume size by the number of pages (~512/volume). In
recent years, however, our condensed format has — for most papers — produced
pages containing far more words than those found in our older volumes, so that it has
taken longer to fill a volume.

Beginning with Mycoraxon 130, we move to the publication of one volume
(issued quarterly) per year. Pagination will run consecutively through all four issues,
but we will continue to list expert reviewers, errata, and nomenclatural novelties
in each quarterly for immediate reference. Although the subscription period now
coincides with the calendar year, those subscribing to our journal only for 2015 will
still be able to access MycoTAxoONn 130 papers in future years for no extra access fee.
Our new ‘nomenclatural’ change brings us in line with other scholarly journals, most
of which also assess annual subscription rates per ‘volume:

We are pleased to announce that with this move, we are FINALLY back on schedule.
The 2015 JANUARY-MARCH quarterly (this issue) presents all papers submitted to
the Editor-in-Chief as of March 31. No longer tied to a targeted number of pages per
volume, we will be able to deliver future quarterlies promptly and on time. Librarians
(and authors) rejoice!

MyYcoTAXON 130(1) contains 37 research papers by 132 authors (representing 19
countries) and revised by 63 expert reviewers.

Within its pages are 32 species and one variety new to science representing
Ambomucor, Anungitea, Bertia, Coccomyces, Ellisembia, Fistulina, Leiorreuma,
Lophodermium, Periconiella, Phyllachora, Rosellinia, Terriera, Tuber, Verticicladus,
and Xylaria from China; Datronia, Hyphoderma, and Lactarius from India;
Dentipellicula from South Africa; Perenniporia from Korea; Pseudoplectania from
Slovakia and the Czech Republic; Sarcoporia from the Canary Islands and the U.S.A.;
Xylaria (again) from Thailand, and Zasmidium from Nepal. We also offer one new
name in Periconiella, one new Cladonia combination (resulting from the taxonomic
clarification of C. corymbescens), and phylogenetic and pathogenicity confirmation of
the synonymy of Bipolaris zizaniae with B. oryzae.

In addition to range extensions for previously named taxa of myxomycetes (for
Turkey and Mexico) and chytrids, mildews, morels, and rusts (for Turkey), we present
a type study of Amauroderma species from China, a key to Chinese Leiorreuma
species, and a complete list of Alternaria species assigned to A. sect. Porri. The
applicability of Distance 1D protein profile analysis for fungi and AFLP analyses in
Coprotus are also evaluated.

Warm regards,

Lorelei L. Norvell (Editor-in-Chief)
13 April 2015

MY COTAXON

http://dx.doi.org/10.5248/130.1
Volume 130, pp. 1-10 January-March 2015

Pseudoplectania lignicola sp. nov.
described from central Europe

S. GLEJDURA™, V. KUCERA?, P. LIZON?, & V. KUNCA!?

'Faculty of Ecology and Environmental Sciences, Technical University in Zvolen,
T. G. Masaryka 24, Zvolen 960 53, Slovakia

?Institute of Botany, Slovak Academy of Sciences,
Dubravska cesta 9, Bratislava 845 23, Slovakia

* CORRESPONDENCE TO: glejdura@gmail.com

ABSTRACT — A new species from Slovakia and the Czech Republic, Pseudoplectania lignicola,
is described and illustrated. It is distinguished from other members of the genus by a centrally
arranged globose membranous sheath surrounding the spores, thick ectal excipulum of
oblong cells at the apothecial base, and growth on less specific biotopes. Comparisons with
similar species and the diagnostic significance of membranous sheath surrounding the
ascospores are also discussed.

Keyworps — biodiversity, Sarcosomataceae, typification

Introduction

The genus Pseudoplectania was established by Fuckel (1870) for two fungi,
Peziza nigrella Pers. and Peziza fulgens Pers. Pseudoplectania nigrella was
selected by Seaver (1913) as the lectotype of the genus, and the species itself
was neotypified by Carbone & Agnello (2012). Peziza fulgens is today treated
in the genus Caloscypha Boud.

Eleven species are currently accepted in the genus: Pseudoplectania affinis,
P. carranzae, P. ericae, RP. kumaonensis, P. melaena, P. nigrella, P. ryvardenii,
P. sphagnophila, P. stygia, PR tasmanica, and P. vogesiaca (Carbone et al. 2013,
2014; Iturriaga et al. 2012).

The first finding of the new Pseudoplectania reported here was associated
with Leucobryum glaucum and tentatively identified in situ as P. sphagnophila.
The new fungus was later collected on Abies alba wood buried in soil as well
as among Sphagnum bogs. The species was mostly collected on conifer wood.

2 ... Glejdura & al.

Materials & methods

The study is based on the morphological examination of 11 collections from three
localities in Slovakia and one in the Czech Republic. The macro-morphological characters
were observed in fresh material. The micro-morphological structures were studied in
fresh and dried material using a light microscope with oil immersion lens (x1000).
Vertical sections of material were examined in 5% KOH, tap-water, Melzer’s reagent,
and a Congo Red + ammonia solution. Values of micro-morphological characters
were estimated as average plus and minus a standard deviation of 30 measurements
for each taxon (with 10 and 90 percentiles presented in parentheses). All descriptions
are based on studied specimens. Localities are georeferenced and the coordinates are
in WGS 84 system. Examined specimens are kept in herbarium BRA, SAV, and HR.
The abbreviations of herbaria are cited in accordance with Index Herbariorum (Thiers
2014).

Taxonomy

Pseudoplectania lignicola Glejdura, V. Kucera, Lizon, Kunca, sp. nov. FIGs 1, 2
MycoBank MB 805097
Differs from Pseudoplectania nigrella by ascospores surrounded by a centrally arranged

globose membranous sheath and by elongated cells in the outer ectal excipulum layer
forming a palisade perpendicular to the surface.

Type: Slovakia, Nizke Tatry Mts., ca. 3 km N of Hiadefl village, 48°50’7.38”N
19°18’39.84’E, alt. 700 m, SW exposure, 20° slope, dystric cambisol, biotite granodiorite,
in a 100-year old managed forest: Picea abies (L.) H. Karst. 77%, Abies alba Mill., 17%,
Larix decidua Mill. 6%; on wood of Abies alba, 21.IV.2010, leg. S. Glejdura (Holotype,
BRA CR-19347).

ErymMo.oey: lignicola (Latin) = living on wood. Most specimens were collected from

wood.
APOTHECIA at first hemispherical, later cup shaped, plane at full maturity,
1-2.7 cm diam, substipitate or sessile. Hymenium smooth, dark brown to
blackish brown. Margin composed of thick-walled cylindrical or slightly
clavate hairs 15-110 um long, 5.5-6.1 um broad, with 1-3 septa and 1.5-2.3 um
thick walls. Receptacle densely tomentose, covered with thick-walled flexuose,
coiled, distantly septate (60-185 um between septa), black-brown to black
hairs, <600 um x 4.6-5.5 um, walls 0.9-1.5 um thick.

ECTAL EXCIPULUM 180-200 um thick, outermost zone 2-4 cells thick, cells
dark brown, thick-walled, subglobose to elongated 20-27 x 14-18 um, inner
layer of subhyaline elongated cells forming a palisade perpendicularly oriented
to the surface, cells 34-92 x 15-29 um.

MEDULLARY EXCIPULUM embedded in a gelatinous matrix, outer layer a
180-200 um thick hyaline textura intricata, cells cylindrical, allantoid, narrowly
pyriform 40-60 x 12-22 um. Inner layer a 500-600 um thick hyaline densely
packed textura intricata, hyphae 9-13 um diam. SUBHYMENIUM 30-40 um

Pseudoplectania lignicola sp. nov. (Slovakia) ... 3

FiGurRE 1. Pseudoplectania lignicola (Holotype BRA CR-19347): A. Ectal excipulum, outermost
thick-walled cells. B. Inner layer of the ectal excipulum, elongated cells perpendicularly arranged
to the surface. C. Distantly arranged cells of medullary excipulum. Scale bar = 50 um.

thick, composed of light brown densely arranged cylindric, globose and
ellipsoid cellular hyphae 3-4.6 um diam.

ASCI narrowly cylindrical, tapered below to thin flexuous aporhynchous
base (simple-septate), apex J-, operculate, 8-spored, 259-299 x 12.3-13.7 um.

4 ... Glejdura & al.

v
o 8

Cc oS

FiGuRE 2. Pseudoplectania lignicola (Holotype BRA CR-19347). A. Ascus with ascospores from
living apothecium; cytoplasmic granules concentrated symmetrically in a limited space by
membranaceous sheath (arrowed). B. Ascospores with centrally arranged membranaceous sheath,
from living apothecium. Scale bar = 10 um.

ASCOSPORES (mature) globose, thick-walled, with granulate or small guttulate
non-oleaginous content (negative reaction in SUDAN 4), (10.2-)11-12.2(-12.3)

Pseudoplectania lignicola sp. nov. (Slovakia) ... 5

um. Mature ascospores surrounded by a membranous cover (sheath) 15.5-19.5
um (including the sheath). The spores are visible in the center of the sheath in
almost all mature ascospores, and in exsiccatae sheath remnants are at least
visible on several spores. Ascospores within membranous sheaths are separated
by cytoplasmic granules laid crosswise in the ascus. Spore walls usually swollen
in 5% KOH, with the membranous sheaths dissolving in the same medium.

PARAPHYSES cylindric, septate, brownish in the upper part and often
agglutinated with an amorphous substance, sometimes branched and
anastomosed in basal and medial parts, 2.2-2.8 um diam, apex mostly straight,
often lobed or diverticulate, with lateral proliferations in upper part, enlarged
to 2.8-3.6 um diam. Irregularly cylindrical paraphyses can be observed in
young or senescent apothecia. Light brown aseptate paraphyses with slightly
thicker walls (so-called hymenial hairs) also present in all stages of apothecial
development.

ECOLOGY & DISTRIBUTION: The species is known from Slovakia and the
Czech Republic and possibly also from Norway (Eckblad 1968, see comment
below). All available collections come from sub-montane and montane moist
coniferous forests (dominated by Picea abies) on acid soils.

ADDITIONAL SPECIMENS EXAMINED: SLOVAKIA, NizkeE Tatry Mrs., ca. 3 km N of
Hiade! village, buffer-zone of Nizke Tatry National Park, 48°50’7.38”N 19°18’39.84’E,
alt. 700 m, in the forest, SW exposition, 20° slope, sporadically rocky, dystric cambisol,
biotite granodiorite, two-storeyed 100-year old production forest, 1“ storey: Picea abies
77%, Abies alba 17%, Larix decidua 6%; 2‘ storey: Fagus sylvatica L. 100%, among moss
Leucobryum glaucum Schimp., 20.I1V.2008, leg. P. Stefanovie (SAV F-10755); on buried
wood of Picea abies, 23.III.2010, leg. S. Glejdura (SAV F-10756); on wood of P. abies,
29.V.2011, leg. S. Glejdura (SAV F-10757); VEPORSKE VRCHY MTs., ca. 1 km SW of
Sihla village, Nature Reserve Vrchslatina, 48°39’18.96”N 19°37’58.50’E, alt. 900 m, in
the forest, flat, 0° slope, dystric histosol (organic soil), diluvial organic sediments (upland
moor) on biotite tonalites and granodiorites, three-storeyed ca 65-year old protective
forest, 1“ storey: Picea abies 100%, 24, 3rd storey: Picea abies 80-90 %, Alnus incana Mill.
20-10%, Sphagno palustris-Piceetum, well-preserved wetland to peatbog biocenoses
with occurrence of specific ecotype of spruce (resonance wood), on buried wood of P.
abies, 28.V.2010, leg. S. Glejdura & V. Kunca (SAV F-10758); on buried wood of P. abies,
leg. S. Glejdura & V. Kunca (SAV F-10759); 2.V1.2010 (SAV F-10760); VYsoKE TAaTRY
Mts., Ticha dolina valley, ca. 6 km NNE of Podbanské village, National Nature Reserve
Ticha dolina, 49°12’1.20’N 19°55’33.24’E, alt. 1140 m, in the forest, NW exposition,
35° slope, typical podzol soil, diluvial sediments on porphyric granitoids and granites,
one-storeyed 55-year old protective forest, Picea abies 65%, Sorbus aucuparia L. 35%, on
dead Sphagnum sp. and needles of P. abies, 18.V.2011, leg. S. Glejdura (SAV F-10761); on
buried wood of Picea abies, among Sphagnum sp. bog, 18.V.2011, leg. S. Glejdura (SAV
F-10762); on cone of Picea abies, 18.V.2011, leg. S. Glejdura (SAV F-10763).

CZECH REPUBLIC, Hraprec KrALove, Orlicka tabule, Chvojnicka plogina,
50°10’45”N 15°54’15”E, alt. 260 m, among moss Leucobryum glaucum in mixed forest
of Picea abies and Pinus sylvestris, 1.1V.2011, leg. Z. Egertova & M. Sochor (HR 89756).

6 ... Glejdura & al.

Discussion

Pseudoplectania is a morphologically and genetically well-delimited genus in
the Sarcosomataceae Kobayasi (Carbone et al. 2013, 2014). Identification of taxa
is based on the paraphysis morphology, ascospore and apothecium size, and
substrate preferences. As we have presented in this paper, another character—
the shape of the gelatinous (mucilaginous) sheath of the spore—must also be
considered an important diagnostic character. Its nature distinguishes similar
species: P. nigrella, P. melaena, and P. lignicola. It is easily detected in fresh
material, less in dry herbarium specimens, although in the type specimen of
P. lignicola the gelatinous sheath was still visible as late as three years after
collection. Spores of P. lignicola lie at the centre of a spherical sheath, whereas
in P. nigrella spores lie to one side of the surrounding sheath (Fics 2, 3). The
hemispherical sheath of spores of P. melaena does not surround the spores but
lies laterally adjacent to the spore surface (Fic 4), a phenomenon that has not
been reported before.

Aside from the spore sheath morphology, Pseudoplectania nigrella (Pers.)
Fuckel differs from P. lignicola by the thickness and textura of the ectal
excipulum as well as by habitat and substrate preferences (on soil in coniferous
forests, rarely on wooden substrates). Also, the ectal excipulum at the base of
its apothecium is thinner (80-100 um) and composed of smaller cells (17-25
um diam) having a textura angularis-globulosa. Eckblad (1968: 119, fig. 65d)
illustrated a specimen of P. nigrella from Norway that has centrally positioned
spores in a membranous sheath. We suspect that this could well represent
P lignicola, so that our new species may have a wider distribution than currently
recorded.

Pseudoplectania melaena (Fr.) Sacc., which has bigger (20-60 mm diam)
stipitate apothecia and apically hooked paraphyses, grows on Abies alba
branches or logs buried in wet soil. Pseudoplectania sphagnophila (Pers.) Kreisel,
associated with Sphagnum sp., has smaller apothecia (10-15 mm diam), and its
paraphyses, which may or may not have lateral branches on the upper parts,
are apically hooked, as in Otidea (Kreisel 1962, Bauer 1999). Pseudoplectania
ericae Donadini, associated with Ericaceae in the Mediterranean area, produces
astipitate apothecia <10 mm diam. and paraphyses that are 2-3-branched at the
apex (Donadini 1987).

Pseudoplectania stygia (Berk. & M.A. Curtis) Sacc. differs mainly by its small
(ca 13 mm diam) apothecia and straight cylindrical paraphyses (Carbone 2013),
while P. ryvardenii Iturr. et al. is distinguished from all other Pseudoplectania
species by its very small (<7 mm diam) apothecia and ornamented ascospores.
Finally, PB kumaonensis Sanwal has distinctly agglutinated paraphyses, and
P. carranzae (Calonge & M. Mata) M. Carbone et al. is separated from other

Pseudoplectania lignicola sp. nov. (Slovakia) ... 7

FiGuRE 3. Pseudoplectania nigrella (SAV F-10764). A. Ascus with ascospores from living apothecium;
cytoplasmic granules concentrated asymmetrically in a limited space by membranaceous sheath
(arrowed). B. Ascospores with eccentrically arranged membranaceous sheath, from living
apothecium. Scale bar = 10 um.

members of the genus mainly by the presence of a tomentum or subicular
hyphae and smaller asci (170-200 x 10-14 um; Calonge & Mata 2002).

8 ... Glejdura & al.

QO;
ao ho

FiGuRE 4. Pseudoplectania melaena (SAV F-10516). A. Ascus with ascospores from living
apothecium; cytoplasmic granules concentrated asymmetrically in a limited space by
membranaceous sheath (arrowed). B. Ascospores with laterally arranged membranaceous sheath,
from living apothecium. Scale bar = 10 um.

Based on our comparison of paraphysis tips in herbarium material of
Pseudoplectania lignicola, P. nigrella, and P. sphagnophila with available literature
data, we conclude that paraphysis shape is not helpful for distinguishing
species. We observed that paraphyses are generally cylindrical, straight, and
unbranched in young apothecia, whereas paraphyses in mature and senescent
apothecia are branched, anastomosed, and often curved at the tip.

Pseudoplectania lignicola sp. nov. (Slovakia) ... 9

The nature of the membranous sheath of the spore, however, is unique
for each Pseudoplectania taxon for which this feature has been observed.
The spores of P lignicola lie at the center of this sheath, and its apothecia are
<27 mm diameter. The new species probably has a wide ecological amplitude,
given its growth on Sphagnum sp., wood, Leucobryum glaucum, and bare soil.
Pseudoplectania species usually occur on the same site for years (when the
conditions allow), and we have registered and collected P lignicola annually
from 2009 to 2013.

In Slovakia P lignicola is rare and possibly endangered (due to lodging and
biotope changes) and should be included in the next edition of the Red List
of Slovak fungi. The fungus has probably been overlooked and incorrectly
identified in the past, but its occurrence in suitable habitats is currently not
seriously threatened.

Acknowledgements

We would like to thank Peter Stefanovie for help in researching the type site of
Pseudoplectania lignicola (Hiadef village), Vaclav Kautman for assistance in the field,
Ginter Bauer for providing material of P. sphagnophila, and Gerard Thijsse and the staff
of the herbarium at Leiden for the opportunity to study Persoon’s herbarium specimens.
Matteo Carbone (Genova, Italy) and Ricardo Galan (Madrid, Spain), who read and
commented on the manuscript, and the support of Shaun Pennycook (Auckland, New
Zealand) are acknowledged. This work was supported by the VEGA [02/0088/13,
02/0150/12] for Kucera and Lizon and by the VEGA [1/1190/12, 1/0362/13] for Glejdura
and Kunca.

Literature cited

Bauer G. 1999. Pseudoplectania sphagnophila (Pers.: Fr.) Kreisel (Ascomycota, Pezizales,
Sarcoscyphaceae) erstmals in Bayern nachgewiesen. Mycol. Bavar. 3: 44—49.

Berkeley MJ. 1875. Notices of North American fungi. Grevillea 3: 145-160.

Calonge FD, Mata M. 2002. Plectania carranzae sp. nov. (Ascomycota) from Costa Rica. Mycotaxon
81: 237-241.

Carbone M. 2013. A type study of Pseudoplectania stygia (Pezizales). Ascomycete.org 5(1): 33-38.

Carbone M, Agnello C. 2012. Studio e tipificazione di Pseudoplectania nigrella. Study and
typification of Pseudoplectania nigrella. Ascomycete.org 4(4): 79-93.

Carbone M, Agnello C, Alvarado P. 2013. Phylogenetic studies in the family Sarcosomataceae
(Ascomycota, Pezizales). Ascomycete.org 5(1): 1-12.

Carbone M, Agnello C, Alvarado P. 2014. Phylogenetic and morphological studies in the genus

Pseudoplectania (Ascomycota, Pezizales). Ascomycete.org 6(1) : 17-33.

Donadini JC. 1987. Etude des Sarcoscyphaceae ss. Le Gal (1). Sarcosomataceae et Sarcoscyphaceae
ss. Korf. Le genre Pseudoplectania emend. nov. P. ericae sp. nov. (Pezizales). Mycol. Helv. 2:
217-246.

Eckblad F-E. 1968. The genera of operculate discomycetes. A re-evaluation of their taxonomy,
phylogeny and nomenclature. Nytt Mag. Bot. 15. 191 p.

Fuckel L. 1870. Symbolae mycologicae. Beitrage zur Kenntnis der rheinischen Pilze. Jahrb. Nass.
Ver. Naturk. 23-24. 459 p.

10 ... Glejdura & al.

Iturriaga T, Mardones M, Urbina H. 2012. A new species of Pseudoplectania (Sarcosomataceae,
Pezizales) from Venezuela. Kurtziana 37(1): 73-78.

Kreisel H. 1962. Pilze der Moore und Ufer Norddeutschlands. III. Pseudoplectania sphagnophila
(Fr. pro var.) Kreisel nov. comb. Westfal. Pilzbriefe 3(5): 74-78.

Saccardo PA. 1889. Sylloge Discomycetum et Phymatosphaeriacearum. Sylloge Fungorum, vol.
8. Padova. 1143 p.Sanwal BD. 1953. Contributions towards our knowledge of the Indian
discomycetes I. Sydowia 7: 191-199.

Seaver FJ. 1913. The genus Pseudoplectania. Mycologia 5(6): 299-302.
http://dx.doi.org/10.2307/3753586

Thiers B. 2014 [continuously updated]. Index Herbariorum: a global directory of public
herbaria and associated staff. New York Botanical Garden's Virtual Herbarium. Available:
http://sweetgum.nybg.org/ih/ [accessed October 2014]

Van Vooren N, Moyne G, Carbone M, Moingeon J-M. 2013. Pseudoplectania melaena (Pezizales):
taxonomical and nomenclatural note. Ascomycete.org 5(1): 47-52.

MYCOTAXON

http://dx.doi.org/10.5248/130.11
Volume 130, pp. 11-15 January-March 2015

Additions to rust and chytrid pathogens of Turkey

CUMALI OZASLAN”, MAKBULE ERDOGDU’,
ELSAD HUSEYIN? & ZEKIYE SULUDERE?

'Dicle University, Faculty of Agriculture, Department of Plant Protection, Diyarbakir-Turkey

?Ahi Evran University, Faculty of Science and Literature, Department of Biology,
Kursehir-Turkey

°Gazi University, Faculty of Science, Department of Biology, Ankara- Turkey

*CORRESPONDENCE TO: cumali.ozaslan@dicle.edu.tr

Asstract — Uromyces bornmuelleri on Bongardia chrysogonum and Physoderma maculare
on Alisma plantago-aquatica have been recently discovered in Turkey. Morphological data
obtained by light and scanning electron microscopy of identified fungi are presented.

Key worps — new records, Chytridiomycota, Basidiomycota

Introduction

The Pucciniaceae are the largest family within the Uredinales, parasitising
almost all major angiosperm orders, with some primarily herbaceous plant
families such as Asteraceae and Poaceae being most commonly infected (van
der Merwe et al. 2007). Uromyces (Link) Unger was raised to generic rank by
Unger (1832), and is typified by U. appendiculatus on Phaseolus. Seventy-four
species of Uromyces have been registered on 227 species of higher plants in
Turkey (Bahcecioglu & Kabaktepe 2012).

All known representatives of the genus Physoderma Wallr.
(Blastocladiomycetes, Physodermataceae) are obligate parasites of vascular
plants (Olson et al. 1980). The generalized life cycle of Physoderma is composed
of two distinct and separate phases, and the ephemeral epibiotic sporangium
with an endobiotic rhizoidal system is confined to a single host cell (Sparrow
& Johns 1965). Physoderma maculare, the type species of the genus, on Alisma
confirms Clinton's (1902) account of the epibiotic stage being produced from
resting zoospores. The latter on mature host tissue may also give rise to the
endobiotic stage, which bears resting spores. On seedlings, however, resting
zoospores produce only epibiotic sporangia (Sparrow 1964).

12 ... Ozaslan & al.

Fics 1-7. Uromyces bornmuelleri. 1: Deformation of host plant. 2: Telia on leaf, in situ. 3: Teliospores.
4-6: Telia and teliospores (SEM). 7. Teliospores (SEM).

Materials & methods

Plant specimens infected with microfungi were collected from Adiyaman and
Siirt provinces, Turkey and prepared according to established herbarium techniques.
Host plants were identified using the Flora of Turkey and East Aegean Islands (Davis
1965-85). The fungal specimens were prepared from the host plants by obtaining thin
sections. Measurements were made from tissues mounted in 5% KOH or tap water.
Microscopical features were examined and microphotographs were made using a Leica

Uromyces and Physoderma spp. new for Turkey... 13

DM E light microscope. Thirty spores were measured for each sample. The microfungi
were identified using relevant literature (De Toni 1888, Saccardo & Sydow 1902,
Gonzalez Fragoso 1918, Kuprevich & Ulijanishchev 1975, van der Merwe et al. 2007).
Species names follow Index Fungorum (2014). The examined specimens have been
deposited in the mycological collection of the Department of Plant Protection, Faculty
of Agriculture, Dicle University, Diyarbakir, Turkey (DUF-M).

For scanning electron microscopy (SEM), 8-10 mm squares of infected leaves were
mounted on SEM stubs with double-sided adhesive tape, coated with gold using a
Polaron SC 502 Sputter Coater, and examined with a Jeol JSM 6060 scanning electron
microscope at 5-10 kV in the Electron Microscopy Unit, Faculty of Science, Gazi
University (Turkey).

Taxonomy

Uromyces bornmuelleri Magnus, Verh. Ges. Deutsch. Naturf. 65: 151.1893. Fics 1-7
SPERMOGONIA, AECIDIA and UREDINIA unknown. TELIA amphigenous,

chestnut brown, in dense groups or scattered, 0.2-3 mm diam., causing leaf
deformation, at first covered by the epidermis, later becoming erumpent,
pulverulent. TELrospores yellow or yellowish-brown, ovoid, globoid,
sometimes oblong, 21-30 x 18-21.5 um, rounded at the apex, rounded or
sometimes attenuate at the base; wall 2-4 um thick, pedicels hyaline, short,
fragile.

SPECIMEN EXAMINED — TURKEY, ADIYAMAN PROVINCE, Center, Kahta, Gerger, Sincik,

Celikhan, Samsat, in wheat field ecosystem, on the leaves of Bongardia chrysogonum (L.)

Spach (Berberidaceae), 12.V.2009, C. Ozaslan CO2029 (DUF-M).
Uromyces bornmuelleri has been reported from Azerbaijan on Leontice
chrysogonum |= Bongardia chrysogonum] (Tranzschel 1939), and from Iraq and
Cyprus on Bongardia chrysogonum (Georghiou 1957, Mathur 1972). Uromyces
bornmuelleri is reported for the first time from Turkey.

Physoderma maculare Wallr., Fl. Crypt. Germ. 2: 192. 1833. Fries 8-13

RESTING SPORANGIA amphigenous, mostly hypophyllous and on leaf
petioles, concentrated on leaves along the nerves, scattered, covered by the
unbroken epidermis, ellipsoid, ovoid, sometimes punctiformis, 1-2 mm diam.,
chestnut brown. RESTING sporEs chestnut brown, globoid, ovoid to ellipsoid,
content granular with more refractive globules, (23.5-)25.5-32 x 21.5-29 um;
with two distinct walls, the inner wall thin and colorless, the outer wall chestnut
brown, 1-2 um thick, smooth. ZoosPorREs not seen.

SPECIMEN EXAMINED — TURKEY, SurtT PROVINCE, Karaca Village, 38°07 09 N
42°01 59 E, in rice field ecosystem, on the leaves of Alisma plantago-aquatica L.
(Alismataceae), 8.V1.2012, C. Ozaslan CO2035 (DUF-M).
The genus Physoderma and P. maculare are reported for the first time from
Turkey. This fungus has been reported from Canada (Conners 1967), Ireland

14 ... Ozaslan & al.

Ae 1 Avr
/
We.

xSBB SE find ie 41-668.

Figs 8-13. Physoderma maculare. 8: Resting sporangia on leaf, in situ. 9: Resting spores. 10: Resting
sporangia on leaf (SEM). 11-13: Resting spores (SEM).

(Muskett & Malone 1984, as Cladochytrium alismatis), Poland (Czeczuga et
al. 2007), Siberia (De Toni 1888, as Uredo alismatis), Spain (Gonzalez Fragoso
1918, as U. alismatis), and U.S.A. (Farr et al. 1989).

Acknowledgements

The author thanks Prof. Dr. Z.M. Azbukina (Vladivostok, Russia) and Dr. Yuri
Tykhonenko (Kiev, Ukraine) for critically reading the manuscript and serving as
presubmission reviewers. Especially we are grateful to Dr. Shaun Pennycook (Auckland,

Uromyces and Physoderma spp. new for Turkey... 15

New Zealand) for detailed linguistic help and nomenclatural review of the manuscript.
We would like thank DUBAP (Dicle University Research Projects Coordinator) for
financial support of this Project (DUBAP 12ZF71). We also thank Prof. Dr. Mecit Vural
(Gazi University, Ankara) and Prof. Dr. A. Selcuk Ertekin (Dicle University, Diyarbakir)
for his help with the identification of the hosts.

Literature cited

Bahcecioglu Z, Kabaktepe $. 2012. Checklist of rust fungi in Turkey. [Mycotaxon 119: 494.]
http://www.mycotaxon.com/resources/checklists/Bahcecioglu-v119-checklist.pdf

Clinton GP. 1902. Cladochytrium alismatis. Botanical Gazette 33: 49-61.

Conners IL. 1967. An annotated index of plant diseases in Canada and fungi recorded on plants
in Alaska, Canada and Greenland. Research Branch, Canada Department of Agriculture,
Publication 1251. 381 p.

Czeczuga B, Muszynska E, Godlewska A, Mazalska B. 2007. Aquatic fungi and straminipilous
organisms on decomposing fragments of wetland plants. Mycologia Balcanica 4: 31-44.

Davis PH (ed.). 1965-85. Flora of Turkey and East Aegean Islands. Vols 1-9. Edinburgh University
Press, Edinburgh.

De Toni GB. 1888. Sylloge ustilaginearum et uredinearum. Sylloge Fungorum 7(2): 449-882.

Farr DF, Bills GF, Chamuris GP, Rossman AY. 1989. Fungi on plants and plant products in the
United States. APS Press, St Paul MN. 1252 p.

Georghiou GP, Papadopoulos C. 1957. A second list of Cyprus fungi. Government of Cyprus,
Department of Agriculture.

Gonzalez Fragoso R. 1918. La roya de los vegetales. Enumeracion y distribucion geografica de los
Uredales. Conocidos hasta hoy en la Peninsula Iberica e Islas Baleares. Trabajos del Museo
Nacional de Ciencias Naturales, Serie Botanica 15: 1-267.

Index Fungorum. 2014. http://www.indexfungorum.org/names/names.asp. Accessed 26 May 2014.

Kuprevich VF, Ulijanishchev V. 1975. Key to the rust fungi in SSSR. Minsk, Belarus: Nauka i
Tekhnika.

Mathur RS. 1972. Checklist of Iraqi Uredinales. Iraq Natural History Museum, Publication 29:
1=32;

Muskett A, Malone J. 1984. Catalogue of Irish fungi - V. Mastigomycotina and Zygomycotina.
Proceedings of the Royal Irish Academy, B. 84: 83-102.

Olson LW, Edén UM, Lange L. 1980. The endobiotic thallus of Physoderma maydis, the causal agent
of Physoderma disease of maize. Protoplasma 103: 1-16.

Saccardo PA, Sydow P. 1902. Supplementum universale, pars V. Sylloge Fungorum 16. 1291 p.

Sparrow FK. 1964. Observations on chytridiaceous parasites of phanerogams. XIII. Physoderma
maculare Wallroth. Archiv fir Mikrobiologie 48: 136-149.

Sparrow FK, Johns RM. 1965. Observation on chytridiaceous parasites of phanerogams XVI.
Notes on Physoderma from Scirpeae. Archiv fiir Mikrobiologie 51: 351-364.

Tranzschel VG. 1939. Compendium of rusts of the USSR. Botanicheskii Institut Akademii Nauk
SSSR. Moscow. 426 p.

Unger F. 1832 [“1833”]. Die Exantheme der Pflanzen. Wien. 422 p.

van der Merwe M, Ericson L, Walker J, Thrall PH, Burdon JJ. 2007. Evolutionary relationships
among species of Puccinia and Uromyces (Pucciniaceae, Uredinales) inferred from protein
coding gene phylogenies. Mycological Research 111: 163-175.

MY COTAXON

http://dx.doi.org/10.5248/130.17
Volume 130, pp. 17-25 January-March 2015

Dentipellicula austroafricana sp. nov. supported by
morphological and phylogenetic analyses

J1A-JIA CHEN*, Lu-LU SHEN* & YU-CHENG DalI*

Institute of Microbiology, Beijing Forestry University, RO. Box 61, Beijing 100083, China
*CORRESPONDENCE TO: daiyucheng2013@gmail.com

ABSTRACT — Dentipellicula austroafricana sp. nov. is described and illustrated from South
Africa based on morphological characters and rDNA sequence data. It is characterized by
an annual growth habit, resupinate basidiocarps, dense soft spines, a monomitic hyphal
structure with non-amyloid, non-dextrinoid and acyanophilous generative hyphae, absence
of cystidia, presence of cystidioles, and tiny rough basidiospores (2.4-2.9 x 2-2.2 um). A
molecular study based on the combined ITS (internal transcribed spacer region) and nLSU
(the large nuclear ribosomal RNA subunit) dataset supports the new species in Dentipellicula.
A key to accepted species of Dentipellicula is provided.

Key worps — Basidiomycota, Hericiaceae, hydnoid fungi, Russulales, taxonomy, wood-
inhabiting fungi

Introduction

Dentipellis Donk (Russulales, Basidiomycota), typified by D. fragilis (Pers.)
Donk, was introduced for species characterized by an annual growth habit,
hydnoid basidiocarps, soft spines, a monomitic hyphal structure with clamp
connections and cyanophilous hyphae, and amyloid rough basidiospores
(Dai et al. 2009, Ginns 1986, Zhou & Dai 2013). Zhou & Dai (2013), who
demonstrated that Dentipellis was polyphyletic, segregated Dentipellis leptodon
(Mont.) Maas Geest. and Dentipellis tatwaniana Sheng H. Wu from Dentipellis
based on ITS and nLSU rDNA sequences. They proposed Dentipellicula Y.C.
Dai & L.W. Zhou as a new genus for these two hydnoid fungal species, which
are distinguished from Dentipellis by their lack of cyanophilous hyphae.
Dentipellicula taiwaniana (Sheng H. Wu) Y.C. Dai & L.W. Zhou (the generic
type), and D. leptodon (Mont.) Y.C. Dai & L.W. Zhou currently comprise the
entire genus.

* Jra-JiA CHEN and Lu-Lu SHEN contributed equally to this work and share first-author status.

18 ... Chen, Shen, & Dai

During studies on the hydnoid fungi, an undescribed Dentipellicula species
from South Africa was identified based on morphological characters and
phylogenetic analysis of ITS and nLSU sequences. We provide an illustrated
description and include an identification key to all three species representing
Dentipellicula.

Materials & methods

Morphological studies

Sections were studied microscopically according to Dai et al. (2010) at
magnifications <1000x using a Nikon Eclipse 80i microscope with phase contrast
illumination. Drawings were made with the aid of a drawing tube. Microscopic
features, measurements, and drawings were made from sections stained with Cotton
Blue and Melzer’s reagent. Spores were measured from sections cut from the tubes. To
present spore size variation, the 5% of measurements excluded from each end of the
range are given in parentheses. Basidiospore spine lengths were not included in the
measurements. Abbreviations include IKI = Melzer’s reagent, IKI- = negative in Melzer’s
reagent, KOH = 5% potassium hydroxide, CB = Cotton Blue, CB+ = cyanophilous,
CB- = acyanophilous, L = mean spore length (arithmetic average of all spores),
W = mean spore width (arithmetic average of all spores), Q = the L/W ratio, and n
= number of spores measured from given number of specimens. Special color terms
follow Petersen (1996). The studied specimens are deposited in the herbaria as cited
below; collection abbreviations follow Thiers (2014).

DNA extraction and sequencing

A CTAB rapid plant genome extraction kit (Aidlab Biotechnologies, Beijing) was
used to obtain PCR products from dried specimens, according to the manufacturer's
instructions with some modifications (Chen & Cui 2014). The DNA was amplified with
the primers: ITS5 and ITS4 for ITS (White et al. 1990), and LROR and LR7 for nLSU
(Vilgalys & Hester 1990). The PCR procedure for ITS was as follows: initial denaturation
at 95°C for 3 min, followed by 35 cycles at 94°C for 40 s, 54°C for 45 s and 72°C for 1
min, and a final extension of 72°C for 10 min. The PCR procedure for nLSU was as
follows: initial denaturation at 94°C for 1 min, followed by 35 cycles at 94°C for 30 s,
50°C for 1 min and 72°C for 1.5 min, and a final extension of 72°C for 10 min. The PCR
products were purified and sequenced in Beijing Genomics Institute, China with the
same primers.

Phylogenetic analysis

New sequences, deposited in GenBank (TaBLE 1), were aligned with additional
sequences retrieved from GenBank (TaBLE 1) using BioEdit (Hall 1999) and ClustalX
(Thompson et al. 1997). Bondarzewia podocarpi Y.C. Dai & B.K. Cui and B. sp. were
used as outgroup (Zhou & Dai 2013). Prior to phylogenetic analysis, ambiguous
regions at the start and the end of the alignment were deleted and gaps were manually
adjusted to optimize the alignment. The edited alignment was deposited at TreeBase
(http://purl.org/phylo/treebase; submission ID 15859).

Dentipellicula austroafricana sp. nov. (China) ... 19

TABLE 1. Specimens used in ITS and nLSU sequence analyses.

GENBANK ACCESSION NO.

SPECIES SAMPLE NO. LOCALITY
ITS nLSU
B. podocarpi Dai 9261 China KJ583207 KJ583221
Bondarzewia sp. DAOM F-415 Canada DQ200923 DQ234539
Dentipellicula austroafricana Dai 12580 South Africa KJ855274 KJ855275
D. leptodon GB 011123 Uganda EU118625 EU118625
D. taiwaniana Dai 10867 China JQ349115 JQ349101
Cui 8346 China JQ349114 JQ349100
Dentipellis coniferarum Cui 10063 China JQ349106 JQ349092
Yuan 5623 China JQ349107 JQ349093
D. dissita NH 6280 Canada AF506386 AF506386
D. fragilis Dai 12550 China JQ349110 JQ349096
Dai 9009 China JQ349108 JQ349094
D. microspora Cui 10035 China JQ349112 JQ349098
D. parmastoi Cui 8513 China JQ349113 JQ349099
Hericium abietis NH 6990 Canada AF506456 AF506456
H. alpestre NH 13240 Russia AF506457 AF506457
H. americanum DAOM F-21467 Canada AF506458 AF506458
H. cirrhatum Tubingen F794 Germany AF506385 AF506385
H. coralloides NH 282 Sweden AF506459 AF506459
H. erinaceus NH 12163 Russia AF506460 AF506460
Laxitextum bicolor NH 5166 Sweden AF310102 AF310102

Phylogenetic analysis followed Li & Cui (2013). Maximum parsimony (MP) analysis
was performed in PAUP* version 4.0b10 (Swofford 2002). All characters were equally
weighted and gaps were treated as missing data. Trees were inferred using the heuristic
search option with TBR branch swapping and 1000 random sequence additions. Max-
trees were set to 5000, branches of zero length were collapsed and all parsimonious trees
were saved. Clade robustness was assessed using a bootstrap (BT) analysis with 1000
replicates (Felsenstein 1985). Descriptive tree statistics tree length (TL), consistency
index (CI), retention index (RI), rescaled consistency index (RC), and homoplasy
index (HI) were calculated for each maximum parsimonious tree (MPT) generated.
Phylogenetic trees were visualized using Treeview (Page 1996).

MrModeltest 2.3 (Nylander 2004) was used to determine the best-fit evolution
model of the combined dataset for Bayesian inference (BI). BI was calculated with
MrBayes 3.1.2 (Ronquist & Huelsenbeck 2003) with a general time reversible (GTR)

20 ... Chen, Shen, & Dai

model of DNA substitution and an invgamma distribution rate variation across sites.
Four Markov chains were performed for 2 runs from random starting trees for 1 million
generations of the combined ITS and nLSU dataset, and trees were sampled every 100
generations. The burn-in was set to discard the first 25% of the trees. A majority rule
consensus tree of all remaining trees was calculated. Nodes that received BT support
>75% and Bayesian posterior probabilities (BPP) 20.95 were considered as significantly
supported.

To determine if the dataset was significantly conflicted, the partition homogeneity
test option in PAUP 4.0b was used between the loci using 1000 replicates and the
heuristic general search option. This test randomly shuffles phylogenetically informative
sites between two paired loci: if the dataset is compatible, shuffling sites between the
loci should not produce summed tree lengths that are significant greater than those
produced by the observed data (Farris et al. 1995; Huelsenbeck et al. 1996).

Molecular phylogeny

Partition homogeneity test showed no conflicts for the two-gene combined
loci (P 20.01). Therefore, in this study, ITS and nLSU were combined into a
single analysis. The combined ITS and nLSU dataset included sequences from
20 fungal collections representing 17 species. The dataset had an aligned length
of 1720 characters, of which 1246 characters are constant, 131 are variable and
parsimony-uninformative, and 343 are parsimony-informative. MP analysis
yielded 2 equally parsimonious trees (TL = 902, CI = 0. 708, RI = 0.786,
RC = 0.557, HI = 0.292). The best model for the combined ITS and nLSU
sequences dataset estimated and applied in the BI was GTR+I+G. BI resulted
in a similar topology with an average standard deviation of split frequencies
= 0.005368 to MP analysis, and thus only the MP tree was provided. Both BT
values (=>50%) and BPPs (20.95) are shown at the nodes (Fic. 1).

The newly sequenced specimen from South Africa was embedded in the
lineage of Dentipellicula (100% MP and 1.00 BPPs) and was closely related to
D. taiwaniana.

Taxonomy

Dentipellicula austroafricana Jia J. Chen, L.L. Shen & Y.C. Dai, sp. nov. Fic. 2
MycoBank MB 811108

Differs from other Dentipellicula species by its exclusively resupinate basidiocarps,
dense spines, absence of gloeoplerous hyphae and cystidia, presence of cystidioles, and
tiny rough basidiospores.

Type — South Africa, KwaZulu-Natal Province, Durban, on charred wood of Ficus,
1.X.2011, Dai 12580 (holotype, BJFC; isotype, IFP; GenBank KJ855274, KJ855275).

ETyMOLoGy — austroafricana (Lat.): referring to South Africa.

FruiTBopy — Basidiocarps annual, resupinate, inseparable, hard corky upon
drying, up to 6 cm long, 2 cm wide, 3 mm thick at center, without odor or taste.

Dentipellicula austroafricana sp. nov. (China) ... 21

87/100 Hericium americanum DAOM F-21467

97/1.00| Hericium alpestre NH 13240

a EF Hericium erinaceus NH 12163
Hericium abietis NH 6990
100/1.00
Hericium cirrhatum Tibingen F794
60/1.00

Hericium coralloides NH 282
Laxitextum bicolor NH 5166
100/0.98| Dentipellis fragilis Dai 12550
99/1.00! ° Dentipellis fragilis Dai 9009

67). Dentipellis dissita NH 6280

Dentipellis parmastoi Cui 8513
100/1.00 | Dentipellis coniferarum Cui 10063

Dentipellis coniferarum Yuan 5623
100/1.00
Dentipellis microspora Cui 10035

86/0.96| Dentipellicula taiwaniana Dai 10867

100/0.99 Dentipellicula tatwaniana Cui 8346

100/1.00 Dentipellicula austroafricana Dai 12580

Dentipellicula leptodon GB 011123
Bondarzewia sp. DAOM F-41

Bondarzewia podocarpi Dai 9261

10
KH

Fic. 1. Strict consensus tree illustrating the phylogenetic position of Dentipellicula austroafricana,
generated by maximum parsimony method based on ITS+nLSU sequence data. Branches are
labeled with parsimony bootstrap values 250% and Bayesian posterior probabilities >0.95.

Fresh spines soft, white to cream, when dry fragile, cream to buff, up to 2 mm
long, 8-10 per mm across base. Margin cottony, buff to clay-buff, <1 mm wide.
Subiculum very thin, soft corky, buff to reddish brown, <1 mm thick.

HyYPHAL STRUCTURE — Hyphal system monomitic; generative hyphae with
clamp connections, IKI-, CB-; tissues unchanged in KOH.

22 ... Chen, Shen, & Dai

SUBICULUM — Generative hyphae colorless, thick-walled, frequently
branched, interwoven, 4-6 um in diam. Gloeoplerous hyphae absent.

HYMENOPHORAL TRAMA — Generative hyphae colorless, slightly thick-
walled to thick-walled, moderately branched, more or less subparallel along
the spines, 2-5.5 um in diam. Gloeoplerous hyphae absent.

HyMENIUM — Cystidia absent; fusoid cystidioles present, hyaline, thin-
walled, 16.5-18 x 1-4 um; basidia clavate with four sterigmata and a basal
clamp connection, 18-23 x 3.5-5 um; basidioles similar to basidia in shape, but
smaller than basidia. Basidiospores ellipsoid to broadly ellipsoid, thick-walled,
colorless, minutely rough, strongly amyloid, CB+, 2.4-2.9(-3) x (1.8-)2-2.2
(-2.4) um, L = 2.83 um, W = 2.12 um, Q = 1.29-1.39 (n = 60/1).

OTHER SPECIES EXAMINED — Dentipellicula leptodon: CANADA. ONTARIO PROVINCE,
Algonquin Nat. Res., on Betula, 22.X.1966 (DAOM F-158439).

Dentipellicula taiwaniana: CHINA. HAINAN PROVINCE, Ledong County, Jianfengling
Nat. Res., on fallen angiosperm trunk, 12.V.2009, Dai 10872 (BJFC 5114).

Key to accepted species of Dentipellicula

PS BasidiOSPOFES = Sior UI LONG cs firs ade deko ech Ase bet lle & wen ke ¥ wereld Berets Baa D. leptodon
Ty BasidiospOrese<t5ro7 UY TOME Ae eta acto Wie doles hte A heated tet dha ee aa 4 2
2. Spities! 5—/. per Mins -Cystidia, presents #4 6.0% Fwhctus Warhol Whee! hee D. austroafricana
2. Spines 8-10 per mm; cystidia absent ................. 00... eee eee D. taiwaniana
Discussion

Phylogenetically, Dentipellicula austroafricana is recognized in the
Dentipellicula clade and distant from Dentipellis, Hericium Pers., and
Laxitextum Lentz based on the combined ITS and nLSU sequences (Fie. 1).
Morphologically, D. austroafricana is characterized by an annual growth habit,
resupinate basidiocarps, soft and dense spines, a monomitic hyphal structure
with non-dextrinoid generative hyphae, presence of cystidioles, and tiny rough
basidiospores. Both morphology and rDNA sequence data confirmed that
D. austroafricana is a new species in Dentipellicula.

According to the ITS and nLSU-based phylogeny (Fic. 1), Dentipellicula
austroafricana is closely related to D. taiwaniana, which also has annual
basidiocarps and soft spines but which is distinguished by its effused-reflexed
basidiocarps, more scattered spines (5-7 per mm), narrower (2-3.5 um)
generative hyphae in the subiculum, presence of gloeocystidia, and longer
basidiospores (2.8-3.4 x 2.1-2.4 um; Dai et al. 2009, Wu 2007).

Dentipellicula leptodon also has annual basidiocarps and soft spines but
differs from D. austroafricana in its narrower (2-4 um) generative hyphae
in the subiculum, presence of gloeopleurous hyphae in the subiculum and
gloeocystidia, and larger basidiospores (3.6-4.2 x 2.8-3.3 um; Dai et al. 2009,

SSS
——

Lak
aa

Wee

QIFR

A
OK ial be
eT"
Go a
\

— f we
Mm I ng a

24 ... Chen, Shen, & Dai

Ginns 1986). Moreover, the two species are separated in the ITS and nLSU-
based phylogenetic analysis (Fie. 1).

Our phylogenetic tree was overall consistent with that presented by Zhou &
Dai (2013): for now, Dentipellis remains polyphyletic based on ITS and nLSU
rDNA sequences, while the new segregate genus Dentipellicula is monophyletic
(Fic. 1). A fully resolved phylogeny for Dentipellis and its related genera requires
evolutionary information from wider taxa samplings and more conserved gene
markers.

Acknowledgements

We express our gratitude to Drs. Li- Wei Zhou (Institute of Applied Ecology, Chinese
Academy of Sciences, China) and Michal TomSovsky (Mendel University in Brno, Czech
Republic) who reviewed the manuscript. The research was financed by the Fundamental
Research Funds for the Central Universities (No. BLYJ201403).

Literature cited

Chen JJ, Cui BK. 2014. Phlebiporia bubalina gen. et. sp. nov. (Meruliaceae, Polyporales) from
southwest China with a preliminary phylogeny based on rDNA sequences. Mycol. Prog. 13:
563-573. http://dx.doi.org/10.1007/s11557-013-0940-4

Cui BK, Zhao CL, Dai YC. 2011. Melanoderma microcarpum gen. et sp. nov. (Basidiomycota) from
China. Mycotaxon 116: 295-302. http://dx.doi.org/10.5248/116.295

Dai YC, Xiong HX, Wu SH. 2009. Notes on Dentipellis (Russulales, Basidiomycota). Mycosystema
28: 668-671.

Dai YC, Cui BK, Liu XY. 2010. Bondarzewia podocarpi, a new and remarkable polypore from
tropical China. Mycologia 102: 881-886. http://dx.doi.org/10.3852/09-050

Farris JS, Kallersj6 M, Kluge AG, Bult C. 1995. Testing significance of incongruence. Cladistics 10:
315-319. http://dx.doi.org/10.1111/j.1096-0031.1994.tb00181.x

Felsenstein J. 1985. Confidence intervals on phylogenetics: an approach using bootstrap. Evolution
39: 783-791. http://dx.doi.org/10.2307/2408678

Ginns J. 1986. The genus Dentipellis (Hericiaceae). Windahlia 16: 35-45.

Hall TA. 1999. BioEdit: a user-friendly biological sequence alignment editor and analysis program
for Windows 95/98/NT. Nucleic Acids Symp. Ser. 41: 95-98.

Huelsenbeck JP, Bull JJ, Cunningham E. 1996. Combining data in phylogenetic analysis. Trends
Ecol. Evol. 11: 152-158. http://dx.doi.org/10.1016/0169-5347(96)10006-9

Li HJ, Cui BK. 2013. Taxonomy and phylogeny of the genus Megasporoporia and its related genera.
Mycologia 105: 368-383. http://dx.doi.org/10.3852/12-114

Nylander JAA. 2004. MrModeltest v2. Program distributed by the author. Evolutionary Biology
Centre, Uppsala University.

Page RDM. 1996. TreeView: application to display phylogenetic trees on personal computers.
Comput. Appl. Biosci. 12: 357-358.

Petersen JH. 1996. The Danish Mycological Societys colour-chart. Foreningen til
Svampekundskabens Fremme, Greve. 6 p.

Ronquist F, Huelsenbeck JP. 2003. MrBayes 3: Bayesian phylogenetic inference under mixed
models. Bioinformatics 19: 1572-1574. http://dx.doi.org/10.1093/bioinformatics/btg180

Swofford DL. 2002. PAUP*: Phylogenetic analysis using parsimony (*and other methods). Version
4.0b10. Sinauer Associates, Sunderland.

Dentipellicula austroafricana sp. nov. (China) ... 25

Thiers, B. 2014 [continuously updated]. Index Herbariorum: A global directory of public herbaria
and associated staff. New York Botanical Garden's Virtual Herbarium.
http://sweetgum.nybg.org/ih/

Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG. 1997. The CLUSTAL_X windows
interface: flexible strategies for multiple sequence alignment aided by quality analysis tools.
Nucleic Acids Res. 25: 4876-4882. http://dx.doi.org/10.1093/nar/25.24.4876

Vilgalys R, Hester M. 1990. Rapid genetic identification and mapping of enzymatically amplified
ribosomal DNA from several Cryptococcus species. J. Bacteriol. 172: 4238-4246.

White TJ, Bruns T, Lee S, Taylor J. 1990. Amplification and direct sequencing of fungal ribosomal
RNA genes for phylogenetics. 315-322, in: MA Innis et al. (eds).PCR Protocols: a guide to
methods and applications. Academic Press, San Diego.

Wu SH. 2007. Three new species of corticioid fungi from Taiwan. Bot. Stud. 48: 325-330.

Zhou LW, Dai YC. 2013. Taxonomy and phylogeny of wood-inhabiting hydnoid species in
Russulales: two new genera, three new species and two new combinations. Mycologia 105:
636-649. http://dx.doi.org/10.3852/12-011

MY COTAXON

http://dx.doi.org/10.5248/130.27
Volume 130, pp. 27-31 January-March 2015

A new species of Terriera (Rhytismatales, Ascomycota)
on Photinia villosa

QinG Lr, YUAN Wu’, DAN-DAN LU’, YA-FEI Xu’, & YING-REN LIN”

" School of Life Science &? School of Forestry & Landscape Architecture,
Anhui Agricultural University, West Changjiang Road 130, Hefei, Anhui 230036, China

*CORRESPONDENCE TO: yingrenlin@yahoo.com

ABSTRACT — A new Terriera species, T: aequabilis on fallen leaves of Photinia villosa from
the Mount Sangingshan National Park, Jiangxi Province, China, is described, illustrated,
and discussed. This taxon is distinguished from T. cladophila by smaller ascomata that are
not associated with conidiomata, black thin zone lines, a well-developed excipulum, and
unbranched paraphyses. The type specimen is deposited in the Reference Collection of Forest
Fungi of Anhui Agricultural University, China (AAUF).

KEY worps — taxonomy, morphology, Rhytismataceae, Rosaceae

Introduction

Eriksson (1970) established the genus Terriera based on T. cladophila
(Lév.) B. Erikss. (© Hysterium cladophilum Lév.), which produces a different
paraphysial type and opening mechanism from genera Lophodermium Cheval.
and Sporomega Corda. Minter (1996) provided a detailed description for
T. cladophila. Johnston (2001), who systematically studied monocotyledon-
inhabiting Lophodermium, divided that heterogeneous genus into five groups,
one of which was transferred to Terriera. IndexFungorum (2014) lists 25
specific and two varietal epithets in Terriera.

Here, Terriera aequabilis on fallen leaves of oriental photinia from Jiangxi
Province, China, is described as a new species.

Materials & methods

Mature ascomata were selected from the collected specimens. Under the dissecting
microscope with a 10—50x magnification, the macroscopic appearance of ascomata and
zone lines was observed. After rehydration of reference materials in water for 15 min.,
vertical transverse 10—15um thick sections were sliced from ascomata using a freezing
microtome and mounted in 0.5% (w/v) cotton blue in water for observing ascomatal

28 ... Li & al.

outlines. Gelatinous sheaths surrounding ascospores and paraphyses were examined
in water or 0.1% (w/v) lactophenol-cotton blue. The color of internal structures and
ascospore contents were observed in water. Squash mounts were prepared in 5% KOH
solution for measurements of asci, ascospores, and paraphyses. Figures of the external
shapes and internal structures of the fruitbodies were drawn using a microscopic
painting tube. The type specimen is deposited in the Reference Collection of Forest
Fungi of Anhui Agricultural University, China (AAUF).

Taxonomy

Terriera aequabilis Qing Li & Y.R. Lin, sp. nov. Fires 1-5
MycoBaAnk MB 808818

Differs from Terriera cladophila by smaller ascomata not associated with conidiomata,
frequent black and thin zone lines, a much thinner covering stroma, unbranched
paraphyses swollen conspicuously at the apex, and ascospores with a visible mucous
sheath.

Type: China, Jiangxi, Mount Sanqingshan National Park, Bingyudong, alt. 1550 m,
on fallen leaves of Photinia villosa (Thunb.) DC. (Rosaceae), 21 August 2012, Y.R. Lin,
S.J. Wang & L. Zhang 2657 (Holotype, AAUF 68765).

ErymMo.oey: aequabilis (Latin = uniform), referring to the single ascospore, which has

an equal diameter throughout its length.
Cotontgs on both sides of leaves, forming distinct subcircular or irregular,
yellow-brown bleached areas each with an obvious edge, 6-10 mm diam.,
which tend to coalesce into larger irregular shapes.

ZONE LINES usually frequent, dark brown or black, thin, entirely or partly
surrounding the paler areas.

CONIDIOMATA not observed.

AscomarTa developing on both sides of leaves, predominantly on the lower
side, scattered over the pale areas, with two ascomata sometimes coalescent. In
surface view, ascomata 320-720 x 250-430 um, elliptical to subcircular, straight
or slightly curved to one side, ends rounded. Whole surface of ascoma black,
except sometimes for a paler region at each end, with a clearly defined edge,
slightly shiny, moderately raising above the substrate surface but somewhat
sunken near the split, opening by a single longitudinal split extending most
of the length of the ascoma. Immature ascomata appearing as a single dark
brown protrusion, more or less rotund in outline, not forming a pale zone
along the future line of opening. In median transverse vertical section,
ascomata subepidermal with epidermal cells becoming filled with fungal tissue
as ascoma develops, 165-190 um deep. COVERING STROMA 15-20 um thick
near the opening, becoming thinner towards the edge and connecting to the
basal stroma, composed mainly of textura angularis with blackish-brown to
dark brown, thick-walled cells 3-5.5 um diam. Lip cells absent. Along the edge
of the ascoma opening, there is a short extension, 10-18um thick, adjacent

Terriera aequabilis sp. nov. (China) ... 29

\\ ASS YTES
SURE F) WISSSEY

ORDO US a St
see OS SoS

Fics 1-5. Terriera aequabilis (holotype, AAUF 68765) on Photinia villosa 1. Habit on a leaf. 2.
Detail of ascomata and a zone line. 3. Ascoma in transverse vertical section. 4. Portion of ascoma
in transverse vertical section. 5. Paraphyses, asci, and ascospores.

30 ... Li&al.

to the top of the covering stroma, which is comprised of strongly carbonized
tissue with no obvious cellular structure. ExcrpuULUM well-developed, 10-15
uum thick, arising from the inner layer of the basal stroma, septate, comprised
of colorless textura porrecta-intricata with hyphae ca 0.8 um diam., sometimes
a reticular structure formed by the interlaced hyphae at the upper part. BASAL
STROMA 6-10 um thick, dark brown, consisting of 2(-3) layers of 3-5 um diam.,
angular, thick-walled cells. Colorless to grayish-brown textura prismatica with
cell 3-8.5 um diam., 20-45 um thick, existing in the triangular area, visible in
sections between the covering and basal stroma at each edge of the ascoma.
SUBHYMENIUM 15-22 um thick, rather flat, consisting of hyaline textura-
angularis and textura intricata. PARAPHYSES 1-1.3 um wide, filiform, sparsely
septate, not branched, gradually or suddenly swollen to 2.8-5 um at the apex,
extending ca 10 um beyond asci, immersed in a ca 0.5 um thick gelatinous
matrix, hyphal bridges no observed at the base. Asc1 maturing sequentially,
cylindric-clavate, 75-105 x 4.5-5.5 um, short-stalked, thin-walled, apex round
or slightly obtuse, J-, 8-spored. Ascosporgs arranged fasciculately, with a
uniform diameter throughout the single ascospore, 55-78 x 0.8-1 um, filiform,
hyaline, aseptate, ends rounded, covered by a 0.3-0.5 um thick gelatinous
sheath.

ADDITIONAL SPECIMENS EXAMINED — On dead leaves of Photinia villosa: CHINA,

J1ANGx1: Mount Sanqingshan National Park, Huaiyushan, alt. ca 600 m, 18 August

2012, S.J. Wang & Y.R. Lin 2632 (AAUF 68740); Sanmucun, alt. ca 850 m, 20 August

2012, F. Zhou & L. Zhang 2645 (AAUF 68753); Yulinfeng, alt. ca 810 m, 21 August 2012,

Y.R. Lin, S.J. Wang & L. Zhang 2704 (AAUF 68812).
ComMMENTsS — Terriera aequabilis is somewhat similar to the type species
T. cladophila in the shape of ascomata, asci, and ascospores. However,
T. cladophila occurs on dead or living twigs and has larger (350-900 x
300-600 um) ascomata that are associated with conidiomata, a much thicker
(<40 um) covering stroma composed of textura globulosa or angularis,
paraphyses with frequently and irregularly branched and irregularly twisted
swollen upper parts, sometimes with hyphal bridges at the base between
adjacent paraphyses, ascospores without mucous sheath, and brown diffuse
zone lines occasionally present (Minter 1996).

Terriera minor (Tehon) P.R. Johnst. [= Lophodermium minus (Tehon) P.R.
Johnst.], the most widely distributed species, differs from the new species in
oblong to oblong-elliptic, paraphyses branching 2-3 times in upper 30-40um,
extend 20-30 um beyond asci and sometimes irregularly swollen at the apices,
longer and wider (100-130 x 6-7 um) asci that are tapering abruptly to small,
rounded apex, wider (1.5-2 um) ascospores slightly tapering towards both
ends and 0-1-septate, and a poorly developed subhymenium (Johnston 1989).
In addition, Tehon’s (1918) observation that a thin, hyaline epithecium was
formed by apically coiled paraphyses differs from Johnston's description.

Terriera aequabilis sp. nov. (China) ... 31

The similar Terriera simplex Y.L. Lin et al. is distinguished from T. aequabilis
by larger (650-1000 x 350-480 um), sometimes triangular ascomata with
conidiomata, a very poorly developed excipulum, wider (1.2-1.5 um)
paraphyses occasionally swollen to 2.5-3 um near the apex, synchronously
maturing asci with truncate to obtuse apices, and ascospores tapering to the
rounded base (Gao et al. 2012).

Acknowledgements

The authors are grateful for the pre-submission comments and suggestions provided
by Dr Z. Wang (Yale University, USA) and Dr M. Ye (Hefei University of Technology,
China) and to F Zhou and L. Zhang for the field investigations. This study was supported
by the National Natural Science Foundation of China (No. 31270065, 31170019).

Literature cited

Eriksson B. 1970. On Ascomycetes on Diapensiales and Ericales in Fennoscandia. I. Discomycetes.
Symbolae Botanicae Upsalienses 19(4): 1-71.

Gao XM, Zheng CT, Lin YR. 2012. Terriera simplex, a new species of Rhytismatales from China.
Mycotaxon 120: 209-213. http://dx.doi.org/10.5248/120.209

IndexFungorum. 2014. [www.indexfungorum.org (viewed online on 1 May 2014)].

Johnston PR. 1989. Lophodermium (Rhytismataceae) on Clusia. Sydowia 41: 170-179.

Johnston PR. 2001. Monograph of the monocotyledon-inhabiting species of Lophodermium.
Mycological Papers 176: 1-239.

Minter DW. 1996. Terriera cladophila. IMI Descriptions of Fungi & Bacteria,. no. 1296.

Tehon LR. 1918. Systematic relationship of Clithris. Botanical Gazette 65: 552-555.
http://dx.doi.org/10.1086/332288

SpeciesFungorum. 2014. [www.speciesfungorum.org (viewed online on 1 May 2014)].

MYCOTAXON

http://dx.doi.org/10.5248/130.33
Volume 130, pp. 33-40 January-March 2015

New records of one Amygdalaria and
three Porpidia taxa (Lecideaceae) from China

Lu-Lu ZHANG, XIN ZHAO, & LING Hu’

Key Laboratory of Plant Stress Research, College of Life Sciences, Shandong Normal University,
Jinan, 250014, P. R. China

* CORRESPONDENCE TO: lichenhuling@gmail.com

AxBstRAcT —Four lichen taxa of Lecideaceae, Amygdalaria consentiens var. consentiens,
Porpidia carlottiana, P. lowiana, and P. tuberculosa, are reported for the first time from China.

Key worps —Asia, Lecideales, lichens

Introduction

The family Lecideaceae Chevall. contains about 23 genera and 547 species,
of which the largest genus is Lecidea Ach., containing about 427 species (Kirk
et al. 2008, Fryday & Hertel 2014). In China, twenty-three species of the other
genera in Lecideaceae have been reported, including two each of Amygdalaria
Norman, Bellemerea Hafellner & Cl. Roux, and Immersaria Rambold &
Pietschm.; one each of Lecidoma Gotth. Schneid. & Hertel, Paraporpidia
Rambold & Pietschm., and Stenhammarella Hertel; and 16 species of Porpidia
Korb. (Hertel 1977, Wei 1991; Aptroot & Seaward 1999; Aptroot 2002; Aptroot
& Sparrius 2003; Obermayer 2004; Guo 2005; Zhang et al. 2010, 2012; Wang et
al. 2012; Ismayi & Abbas 2013; Hu et al. 2014).

Amygdalaria and Porpidia are obviously very closely related. Both have
large halonate ascospores, a high hymenium, Porpidia-type asci, and a dark
pigmented hypothecium and are (with a few exceptions) restricted to lime-free,
silicate rocks. However Amygdalaria can be best distinguished from Porpidia
by the presence of cephalodia, the higher hymenium (over 130 um), the larger
ascospores (generally 20-35 x 10-16 um) with conspicuous, rather compact
epispores, and a tendency toward a brownish or yellowish pink thallus (Inoue
1984, Brodo & Hertel 1987, Gowan 1989, Smith et al. 2009).

34 ... Zhang, Zhao, & Hu

During our research on the lecideoid taxa of Lecideaceae in China, we
have identified four taxa new to the country: Amygdalaria consentiens vat.
consentiens, Porpidia carlottiana, P. lowiana, and P. tuberculosa.

Materials & methods

The studied specimens are preserved in the Lichen Section of Botanical Herbarium,
Shandong Normal University, Jinan, China (SDNU); Herbarium Mycologicum
Academiae Sinicae - Lichenes, Beijing, China (HMAS-L); and the Herbarium of
the Kunming Institute of Botany, Chinese Academy of Sciences, Kunming, China
(KUN). The morphological and anatomical characters were examined under a stereo-
microscope (COIC XTL7045B2) and a polarizing microscope (OLyMpus CX41). The
thalli and medullae were tested for identification with K (10% aqueous KOH solution),
C (saturated aqueous NaClO solution), I (a 10% aqueous KI solution), P (a saturated
p-phenylenediamine solution in 95% ethyl alcohol). The lichen substances were
identified using standardized thin layer chromatography techniques (TLC) with solvent
system C (Orange et al. 2010). Photos of these lichens were taken under Olympus
SZX16 and BX61 with DP72.

Taxonomic descriptions

Amygdalaria consentiens (Nyl.) Hertel, Brodo & Mas. Inoue, J. Hattori Bot. Lab. 56:
322 (1984) var. consentiens Pret

MorPHoLocy — THALLUS continuous, cracked-areolate, usually thick,
light orange to brownish gray, often becoming oxidated orange; medulla I-;
prothallus black, thin and conspicuous; cephalodia mostly flush with thallus
surface, almost the same colour as the thallus, but occasionally pinker or gray,
containing Stigonema; soredia absent. APOTHECIA entirely sunken in thallus,
(0.35-)0.45-0.9 mm diam., usually one or two per areole; disc dark brown to
black, margin slightly to moderately rusty orange pruinose. Excipie black-
brown, 50-70 um wide, without crystals; epihymenium brownish, with orange
crystals; hymenium hyaline, 150-200 um tall; subhymenium colorless, 45-60
um tall; hypothecium blackish brown, usually thin, bowl-shaped; paraphyses
strongly anastomosed and branched, almost moniliform at the apical part,
with +swollen apices. Asci clavate, Porpidia-type; ascospores hyaline, simple,
ellipsoid, 28-32(-35) x 14-17(-19) um, halonate. Pycnip1A immersed, conidia
bacilliform, 6-9 x c. 1 um.

CHEMISTRY — Thallus and medulla K-, C-, KC-, P-. No lichen substances
were detected by TLC.

DISTRIBUTION — Amygdalaria consentiens has been reported from Asia,
Europe, and North America (Inoue 1984, Brodo & Hertel 1987, Smith et al.
2009). New to China.

SPECIMEN EXAMINED: CHINA. YUNNAN, Luquan, Mt. Jiaozixueshan, alt. 3800 m, on
rock, 26 Oct. 2008, Z.J. Ren 20108488 (SDNU).

Amygdalaria & Porpidia species new to China... 35

Fic. 1 Amygdalaria consentiens var. consentiens (Ren 20108488, SDNU). A: thallus; B: prothallus;
C: apothecium section; D: epihymenium with crystals; E: amyloid reaction of ascus; F: ascus and
ascospores; G: ascospores; H: paraphyses.

COMMENTS —Amygdalaria consentiens var. consentiens is morphologically
similar to A. consentiens var. japonica and A. continua, but A. consentiens var.
japonica contains stictic and constictic acids, while A. continua has a smooth to
finely rimose thallus which is rarely cracked-areolate.

Porpidia carlottiana Gowan, Bryologist 92: 39 (1989) Fia. 2
MorPHoLocy — THALLUS crustose, finely cracked to subrimose, 0.15-0.25
mm thick, whitish gray; medulla I-; prothallus continuous between thallus

36 ... Zhang, Zhao, & Hu

20 pm 20 pm

Eb ad

Fic. 2 Porpidia carlottiana (Wang 20127123, SDNU). A: thallus; B: apothecium section; C: amyloid
reaction of ascus; D: ascospores; E: exciple and epihymenium without crystals.

patches, black, thin; soredia absent. APOTHECIA scattered or contiguous,
sunken in thallus when young, becoming sessile on a broad base when mature,
0.5-1.1(-1.5) mm diam.; disc black, plane to weakly convex, usually with
heavy whitish pruina; margin bare, distinct, regular to flexuose. ExcrpLe dark
brown to black at margin, brown within, 90-105 um wide, without crystals;
epihymenium yellowish brown or olive brown, without crystals; hymenium
hyaline, 90-105(-130) um tall; subhymenium 20-25 um, hypothecium
blackish brown; paraphyses strongly anastomosed and apically branched. Asci
clavate, Porpidia-type; ascospores hyaline, simple, ellipsoid, 17-20 x 7-10 um,
halonate. Pycnip1a not observed.

CHEMISTRY — ‘Thallus and medulla K-, C-, KC-, P-. 2’-O-methyl-
superphyllinic acid was detected by TLC.

DISTRIBUTION — Porpidia carlottiana has been reported from North
America (Gowan 1989). New to China.

SPECIMEN EXAMINED: CHINA. YUNNAN, Lijiang, Mt. Laojunshan, alt. 3800 m, on rock,

5 Nov. 2009, H.Y. Wang 20127123 (SDNU).
ComMENts — Porpidia carlottiana is closely related to P. rugosa, which also
produces pruinose apothecia and 2’-O-methylsuperphyllinic acid. However,
P. rugosa always has soredia, and the apothecia rarely seen.

Amygdalaria & Porpidia species new to China... 37

Porpidia lowiana Gowan, Bryologist 92: 49 (1989) Fic. 3

MorPHOoLocy — THALLUS crustose, continuous, finely cracked to rimose-
areolate, 0.2-0.5 mm thick, yellowish gray to whitish, sometimes dark gray
or greenish gray; medulla I-; prothallus absent; soredia absent. APOTHECIA
scattered to contiguous, soon becoming broadly sessile, 0.5-1.3 mm diam.;
disc black, non-pruinose to moderately pruinose; margin bare, distinct,
even to weakly crenulate, regular to flexuose. ExcrpLe mostly black and
carbonaceous at exciple margin, hyaline within, 55-90 um wide, with crystals
that are soluble in K and C; epihymenium olive brown, without crystals;
hymenium hyaline, 70-110 um tall; subhymenium 18-26 um, hypothecium
blackish brown; paraphyses strongly anastomosed and apically branched. Asci
clavate, Porpidia-type; ascospores hyaline, simple, ellipsoid or tear-drop-shape,
14-16(-25) x (4.5-)6-8 um, halonate. Pycnip1A not observed.

CHEMISTRY — ‘Thallus and medulla K-, C-, KC-, P-. Confluentic,
2’-O-methylmicrophyllinic, and 2’-O-methylperlatolic acids were detected by
TLE.

DISTRIBUTION — Porpidia lowiana has been reported from Europe and
North America (Gowan 1989, Smith et al. 2009). New to China.

Fic. 3 Porpidia lowiana (Ren 20102097, SDNU). A: thallus; B: apothecium section; C: amyloid
reaction of ascus; D: ascospores; E: exciple with crystals.

38 ... Zhang, Zhao, & Hu

SPECIMENS EXAMINED: CHINA. HELONGJIANG, Tahe, Mt. Mengkeshan, alt. 555 m, on
rock, 14 Aug. 2009, Q. Ren 20102097 (SDNU). INNER Monco ia, Mangui, alt. 900 m,
on rock, 13 Sep. 1977, J.C. Wei, 074581 (HMAS-L); SicHUAN, Dukou, Mt. Dabaoding,
alt. 1950 m, on rock, 21 Jun. 1983, L.S. Wang 83-184 (KUN); YUNNAN, Xinping, Mt.
Mopan, alt. 2000 m, on rock, 20 Dec. 2008, L.S. Wang 08-30003 (KUN); Chuxiong, Mt.
Zixi, alt. 2060 m, on rock, 31 Aug. 2005, L.S. Wang 05-25263 (KUN); Lincang county,
Matai village, alt. 1650 m, on rock, 20 Oct. 2003, L.S. Wang 03-22921, 03-22922 (KUN);
Yimen, alt. 1600 m, on rock, 10 Dec. 1990, L.S. Wang 11707 (KUN).
ComMENts — Porpidia lowiana is closely related to P cinereoatra, from
which it differs mainly in having a thinner, yellower thallus, thicker proper
margin (c. 0.05 mm in P. cinereoatra), and sessile apothecia (mainly sunken in
P. cinereoatra).

Porpidia tuberculosa (Sm.) Hertel & Knoph, Beih. Nova Hedwigia 79: 438 (1984)
Fic. 4

MorPHOLOGY — THALLUS crustose, continuous, subrimose to distinctly
rimose-areolate, 0.2-0.3(-0.5) mm thick, medium gray or sometimes rusty
orange; medulla I+ violet; prothallus usually present when abutting another
lichen, black; soralia present, 0.3-0.7 mm diam., round to irregular, scattered
to crowded, shallowly erose or abraded, usually with slightly raised rim;
soredia farinose to granular, white, pale gray or flecked blue-gray or blackish,
I+ violet. APOTHECIA occasional, scattered, soon becoming sessile, 0.9-1.5
(-1.7) mm diam.; disc black, plane to subconvex, weakly to moderately pruinose;
margin bare, distinct, regular to flexuose. ExcIPLE greenish to brownish black
at exciple margin, dark brown within, 90-140 um wide, without crystals;
epihymenium olive brown, without crystals; hymenium hyaline, 90-105
um tall; subhymenium 35-50 um, hypothecium blackish brown; paraphyses
strongly anastomosed and apically branched. Ascr clavate, Porpidia-type;
ascospores hyaline, simple, ellipsoid, 17.5-22 x 7-8 um, halonate. PycNnip1A
immersed, conidia bacilliform, 7.5-10 x c. 1 um.

CHEMISTRY — ‘Thallus and medulla K-, C-, KC-, P-. Confluentic,
2’-O-methylmicrophyllinic, and 2’-O-methylperlatolic acids were detected by
TES

DISTRIBUTION — Porpidia tuberculosa has been reported from Asia,
Macaronesia, Europe, and North & South America (Hertel 1984, Smith et al.
2009). New to China.

SPECIMENS EXAMINED: CHINA. HusBEI, Shennongjia, Mt. Dashennongijia, alt. 3000 m,
on rock, 27 Jun. 1984, J.C. Wei, 067994, 068005, 074292 (HMAS-L). YUNNAN, Luquan,
Mt. Jiaozixueshan, alt. 3800 m, on rock, 27 Oct. 2008, H.Y. Wang 20081180, 20082302
(SDNU).
ComMENtTs — Porpidia tuberculosa, P. rugosa and P. soredizodes all have
a thallus with soralia. However, P rugosa has a more verrucose thallus,

Amygdalaria & Porpidia species new to China... 39

100 um

Fic. 4 Porpidia tuberculosa (Wei 067994, HMAS-L). A: thallus; B: apothecium section; C: amyloid
reaction of ascus; D: conidia; E: exciple and epihymenium without crystals.

I- medulla, and contains 2’-O-methylsuperphyllinic and glaucophaeic acids.
Porpidia soredizodes has an I- medulla as well as an overall smaller thinner
(<0.2 mm) darker thallus that contains stictic acid.

Acknowledgements

The authors thank Dr. A. Aptroot (ABL Herbarium, Soest, the Netherlands) and
Dr. Shou-Yu Guo (State Key Laboratory of Mycology, Institute of Microbiology,
Chinese Academy of Sciences, Beijing, China) for presubmission reviews. This study
was supported by Program for Scientific Research Innovation Team in Colleges and
Universities of Shandong Province, the National Natural Science Foundation of China
(31170187), Foundation of Key Laboratory, CAS (KLBB-201306), and the Scientific
Research Foundation of Graduate School of Shandong Normal University (BCX1406).

Literature cited

Aptroot A. 2002. Corticolous and saxicolous lichens from Xishuangbanna, southern Yunnan,
China. http://www.nhm.uio.no/botanisk/lav/Yunnan.

Aptroot A, Seaward MRD. 1999. Annotated checklist of Hong Kong lichens. Tropical Bryology 17:
57-101.

Aptroot A, Sparrius LB. 2003. New microlichens from Taiwan. Fungal Diversity 14: 1-50.

AO ... Zhang, Zhao, & Hu

Brodo IM, Hertel H. 1987. The lichen genus Amygdalaria (Porpidiaceae) in North America.
Herzogia 7: 493-521

Fryday AM, Hertel H. 2014. A contribution to the family Lecideaceae s. lat. (Lecanoromycetidae inc.
sed., lichenized Ascomycota) in the southern subpolar region; including eight new species and
some revised generic circumscriptions. Lichenologist 46(3): 389-412.
http://dx.doi.org/10.1017/S0024282913000704

Gowan SP. 1989. The lichen genus Porpidia (Porpidiaceae) in North America. Bryologist 92: 25-59.

Guo SY. 2005. Lichens. 31-82, in: WY Zhuang (ed.). Fungi of northwestern China. Mycotaxon Ltd.,
Ithaca, New York.

Hertel H. 1977. Gesteinsbewohnende Arten der Sammelgattung Lecidea (Lichenes) aus Zentral-,
Ost- und Siidasien. Khumbu Himal, Ergebnisse des Forschungsunternehmens Nepal Himalaya.
6: 145-378.

Hertel H. 1984. Uber saxicole, lecideoide Flechten der Subantarktis. Beihefte zur Nova Hedwigia.
79: 399-499.

Hu L, Zhao X, Sun LY, Zhao ZT, Zhang LL. 2014. Four lecideoid lichens new to China. Mycotaxon
128: 83-91. http://dx.doi.org/10.5248/128

Inoue M. 1984. Japanese crustose lichen genera formerly reported under Lecidea sensu lato. 1.
Amygdalaria Norman. Journal of the Hattori Botanical Laboratory 56: 321-330.

Ismayi G, Abbas A. 2013. Bellemerea alpina — new lichen species record from China. Plant Science
Journal 31(4): 333-335.

Kirk PM, Cannon PF, Minter DW, Stalpers JA. 2008. Dictionary of the fungi. 10th edition. CABI
Bioscience: CAB International. 771 p.

Obermayer W. 2004. Additions to the lichen flora of the Tibetan region. Bibliotheca Lichenologica
88: 479-526.

Orange A, James PW, White FJ. 2010. Microchemical methods for the identification of lichens. 2nd
edition. London: British Lichen Society.

Smith CW, Aptroot A, Coppins BJ, Fletcher A, Gilbert OL, James PW, Wolseley PA (eds). 2009. The
lichens of Great Britain and Ireland. Natural History Museum Publications, in association with
The British Lichen Society.

Wang XY, Zhang LL, Joshi Y, Wang HY, Hur JS. 2012. New species and new records of the
lichen genus Porpidia (Lecideaceae) from western China. Lichenologist 44(5): 619-624.
http://dx.doi.org/10.1017/S0024282912000242

Wei JC. 1991. An enumeration of lichens in China. International Academic Publishers, Beijing.

Zhang LL, Wang HY, Sun LY, Zhao ZT. 2010. Four lichens of the genus Lecidea from China.
Mycotaxon 112: 445-450. http://dx.doi.org/10.5248/112.445

Zhang LL, Wang LS, Wang HY, Zhao ZT. 2012. Four new records of lecideoid lichens from China.
Mycotaxon 119: 445-451. http://dx.doi.org/10.5248/119.445

MYCOTAXON

http://dx.doi.org/10.5248/130.41
Volume 130, pp. 41-46 January-March 2015

Anungitea guangxiensis and Ellisembia longchiensis,
two new species from southern China

JI-WEN XIA, YING-RuI Ma, & XIU-GUO ZHANG

Department of Plant Pathology, Shandong Agricultural University, Taian, 271018, China

*CORRESPONDENCE TO: zhxg@sdau.edu.cn, sdau613@163.com

ABSTRACT — Two new hyphomycete species, Anungitea guangxiensis and Ellisembia
longchiensis, are described and illustrated from specimens collected on dead branches in
southern China. Anungitea guangxiensis is characterized by conidiogenous cells that are
polyblastic, integrated, terminal, smooth, and denticulate (with cylindrical denticles) and
conidia produced in acropetal chains. Ellisembia longchiensis is characterized by distinct,
single, unbranched, erect conidiophores, monoblastic conidiogenous cells, and distoseptate
conidia that have a small, globose, mucilaginous apical sheath.

Key worps — conidial fungi, taxonomy

Introduction

Anungitea was established by Sutton (1973) as a monotypic genus with
A. fragilis B. Sutton as the type species. The genus is characterized by conidia borne
on conspicuous denticulate loci of the terminal or intercalary conidiogenous
cells and which form acropetal, unbranched chains. A further 19 names have
been proposed in Anungitea (Castafeda 1986; Castafieda & Kendrick 1990,
1991; Castaneda et al. 1997; Crous et al. 1995, 2014; Kirk 1983; Matsushima
1975; Rambelli et al. 2008, 2009, 2010; Sutton & Hodges 1978), but subsequently
A. globosa B. Sutton & Hodges was synonymised with A. fragilis (Kirk 1982;
Matsushima 1987; Castaneda Ruiz et al. 1996), and A. triseptata R.F. Castafieda
& W.B. Kendr. was transferred to Anungitopsis (Castaheda & Kendrick 1991).

The genus Ellisembia was introduced by Subramanian (1992) to accommodate
Sporidesmium-like species with determinate or irregularly percurrently
extending conidiogenous cells that produce distoseptate conidia. Wu & Zhuang
(2005) merged Imicles Shoemaker & Hambl. (Shoemaker & Hambleton 2001)
into Ellisembia and expanded the generic concept to include species that
produce typically lageniform, ovoid, or doliiform percurrently extending
conidiogenous cells.

42 ... Xia, Ma, & Zhang

-—-«.
.
+. a &
.. >
iv.

Peal fe
. -
*

gue
ad 6

———-.
.
_
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& . =
*
al os
“& BLOM a. ee +
lsat Ye
ae = + ae

|
.

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IF

Fic. 1. Anungitea guangxiensis (holotype, HSAUP H6351).
A,C. Conidiophores with conidia. B. Apex of conidiophore.
D. Conidia. E,F. Apex of conidiophores with conidia.

Many taxa of hyphomycetes have been already described from China (Wu
& Zhuang 2005, Zhang et al. 2009a,b, 2011, 2012, Ma et al. 2010, 2012a,b,c, Ren

Anungitea & Ellisembia spp. nov. (China) ... 43

et al. 2012). In our studies on hyphomycetes from deciduous stems and rotten
wood in southern China, we collected two hyphomycetes that possess the
morphological characteristics of Anungitea and Ellisembia. Both are proposed
herein as new species.

Anungitea guangxiensis J.W. Xia & X.G. Zhang, sp. nov. FIG. 1
MycoBank MB 811157
Differs from Anungitea heterospora by its much larger conidia; and from A. longicatenata
by its longer, cylindrical conidia.
Type: China, Guangxi Province: Mount Dayao, on dead stems of unidentified broad-
leaved tree, 26 Oct 2012, J.W. Xia (Holotype, HSAUP H6351; isotype, HMAS 243458).

EryMo_oey: in reference to the province where the type was found.

Colonies on natural substrate effuse, pale brown to brown, hairy. Mycelium
partly superficial, partly immersed in the substratum, composed of septate,
pale brown, smooth, 1-2 um wide hyphae. Conidiophores distinct, single,
unbranched, erect, straight or slightly flexuous, cylindrical, smooth, thick-
walled, pale brown to brown, 6-11-septate, 70-160 x 2.5-4.5 um. Conidiogenous
cells polyblastic, integrated, terminal, sympodial, hyaline to pale brown, 10-18 x
2.5-4.5 um, with 1-6 short denticles. Conidia acrogenous, cylindrical, in
unbranched acropetal chains, dry, subhyaline to hyaline, 1-septate, truncate at
the base and rounded at the apex, 16-22 um long, 3-4 um wide.

ComMENTs - Among the known species, Anungitea guangxiensis bears some
resemblance to A. heterospora P.M. Kirk and A. longicatenata Matsush. in
conidial shape. However, the primary conidia of A. heterospora are smaller
(8-9.5 x 3-4 um, l-septate) than those of A. guangxiensis and occasionally
slightly constricted at the septum (Kirk 1983). The conidia of A. longicatenata
differ by being shorter (11-18 x 2.5-4 um, 0-2-septate), narrowly obovoid to
cylindrical, and tapering at both ends (Matsushima 1975).

Ellisembia longchiensis J.W. Xia & X.G. Zhang, sp. nov. Fic. 2
MycoBank MB 811159
Differs from Ellisembia mucicola by its longer, narrower, obclavate conidia with more

numerous distosepta; and from E. suttonii by its longer, narrower conidia and its
conidiogenous cells lacking percurrent proliferations.

Type: China, Sichuan Province: national forest park of Longchi, on dead stems of
unidentified broad-leaved tree, 17 Apr 2012, J.W. Xia (Holotype, HSAUP H6246-1;
isotype, HMAS 243459).

EryMo_oey: in reference to the type locality.
Colonies on natural substrate effuse, pale brown to brown, hairy. Mycelium
partly superficial, partly immersed in the substratum, composed of septate, pale
brown, smooth, 2-5 um wide hyphae. Conidiophores distinct, single, erect,

44 ... Xia, Ma, & Zhang

A : B
20um

14> — 1 eo

aed .

(Ory) Gs
te

Fic. 2. Ellisembia longchiensis (holotype, HSAUP H6246-1).
A. Conidiophores with conidia. B. Conidia.

straight or slightly flexuous, cylindrical, smooth, thick-walled, pale brown
to brown, 2-4-septate, 55-75 x 3.5-5 um. Conidiogenous cells monoblastic,
integrated, terminal, pale brown to brown, 13.5-15 x 3.5-5 um. Conidial

Anungitea & Ellisembia spp. nov. (China) ... 45

secession schizolytic. Conidia holoblastic, solitary, acrogenous, straight or
slightly curved, obclavate, smooth-walled, pale brown to brown, paler towards
the apex where it is hyaline, 9-12-distoseptate, 55-86 um long, 6-7 um thick
in the broadest part, 3-4.5 um wide at the truncate base, with a small, globose,
mucilaginous apical sheath.

Comments - Ellisembia longchiensis is closely related to E. mucicola W.P. Wu
and E. suttonii W.P. Wu. Ellisembia mucicola differs by its shorter, wider,
ellipsoid to fusiform conidia (42-52 x 2.5-4 um, 8-9-septate); and E. suttonii
differs by its shorter, wider conidia (45-52 x 9-12 um, 10-12-septate) and its
percurrently proliferating conidiogenous cells (Wu & Zhuang 2005).

Acknowledgments

The authors express gratitude to Dr. Eric H.C. McKenzie and Dr. Bryce Kendrick for
serving as pre-submission reviewers and for their valuable comments and suggestions.
This project was supported by the National Natural Science Foundation of China (Nos.
31093440, 31230001) and the Ministry of Science and Technology of the People’s
Republic of China (Nos. 2006FY120100).

Literature cited

Castaneda Ruiz RE 1986. Fungi cubense. Instituto de Investigaciones Fundamentales en Agricultura
Tropical “Alejandro de Humboldt’, C. Habana. 20 p.

Castaneda Ruiz RE, Kendrick WB. 1990. Conidial fungi from Cuba: II. University of Waterloo
Biology Series 33: 1-61.

Castafieda Ruiz RE, Kendrick WB. 1991. Ninety-nine conidial fungi from Cuba and three from
Canada. University of Waterloo Biology Series 35: 1-132.

Castafieda Ruiz RF, Gené J, Guarro J. 1996. Litter hyphomycetes from La Gomera (Canaries).
Mycotaxon 59: 203-215.

Castaneda Ruiz RF, Kendrick WB, Guarro J. 1997. Notes on conidial fungi. XIV. New hyphomycetes
from Cuba. Mycotaxon 65: 93-105.

Crous PW, Wingfield MJ, Kendrick WB. 1995. Foliicolous dematiaceous hyphomycetes from
Syzygium cordatum. Can. J. Bot. 73: 224-234.

Crous PW, Groenewald JZ, Shivas RG. 2014. Fungal Planet 222 - 10 June 2014. Anungitea
eucalyptorum Crous & R.G. Shivas, sp. nov. Persoonia 32: 198-199.

Kirk PM. 1982. New or interesting microfungi. IV. Dematiaceous hyphomycetes from Devon.
Transactions of the British Mycological Society 78: 55-74.

Kirk PM. 1983. New or interesting microfungi. IX. Dematiaceous hyphomycetes from Esher
Common. Transactions of the British Mycological Society 80: 449-467.

Ma LG, Ma J, Zhang YD, Zhang XG. 2010. A new species of Spadicoides from Yunnan, China.
Mycotaxon 113: 255-258. http://dx.doi.org/10.5248/113.255

Ma LG, Ma J, Zhang YD, Zhang XG. 2012a. Spadicoides camelliae and Diplococcium livistonae, two
new hyphomycetes on dead branches from Fujian Province, China. Mycoscience 53: 25-30.
http://dx.doi.org/10.1007/s10267-011-0138-z

Ma J, Zhang YD, Ma LG, Ren SC, Castafieda Ruiz RE, Zhang XG. 2012b. Three new
species of Solicorynespora from Hainan, China. Mycological Progress 11: 639-645.
http://dx.doi.org/10.1007/s11557-011-0775-9

46 ... Xia, Ma, & Zhang

MaJ, Zhang YD, Ma LG, Castaneda Ruiz RE, Zhang XG. 2012c. Three new species of Sporidesmiella
from southern China. Mycoscience 53: 187-193. http://dx.doi.org/10.1007/s10267-011-0152-1

Matsushima T. 1975. Icones microfungorum a Matsushima lectorum. Matsushima, Kobe. 209 p.

Matsushima T. 1987. Matsushima Mycological Memoirs 5: 1-100.

Rambelli A, Venturella G, Ciccarone C. 2008. Dematiaceous hyphomycetes from Pantelleria
Mediterranean maquis litter. Flora Mediterranea 18: 441-467.

Rambelli A, Venturella G, Ciccarone C. 2009. More dematiaceous hyphomycetes from Pantelleria
Mediterranean maquis litter. Flora Mediterranea 19: 81-113.

Rambelli A, Tempesta S, Venturella G, Ciccarone C. 2010. Dematiaceous hyphomycetes from
Pantelleria Mediterranean maquis litter. Flora Mediterranea 20: 211-233.

Ren SC, Ma J, Ma LG, Zhang YD, Zhang XG. 2012. Sativumoides and Cladosporiopsis,
two new genera of hyphomycetes from China. Mycological Progress 11: 443-448.
http://dx.doi.org/10.1007/s11557-011-0759-9

Shoemaker RA, Hambleton S. 2001. “Helminthosporium” asterinum, Polydesmus elegans, Imimyces,
and allies. Canadian Journal of Botany 79: 592-599. http://dx.doi.org/10.1139/cjb-79-5-592.

Subramanian CV. 1992. A reassessment of Sporidesmium (hyphomycetes) and some related taxa.
Proceeding of the Indian National Science Academy, B 58: 179-190.

Sutton BC. 1973. Hyphomycetes from Manitoba and Saskatchewan, Canada. Mycological Papers
132: 1-143.

Sutton BC, Hodges CS. 1978. Eucalyptus microfungi. Chaetendophragmiopsis gen. nov. and other
hyphomycetes. Nova Hedwigia 29: 593-607.

Wu WP, Zhuang WY. 2005. Sporidesmium, Endophragmiella and related genera from China. Fungal
Diversity Research Series 15: 1-351.

Zhang K, Ma J, Wang Y, Zhang XG. 2009a. Three new species of Piricaudiopsis from southern
China. Mycologia 101: 417-422. http://dx.doi.org/10.3852/08-147

Zhang K, Ma LG, Zhang XG. 2009b. New species and records of Shrungabeeja from southern
China. Mycologia 101: 573-578. http://dx.doi.org/10.3852/09-006

Zhang YD, Ma J, Ma LG, Castafieda Ruiz RF, Zhang XG. 2011. New species of Phaeodactylium and
Neosporidesmium from China. Sydowia 63: 125-130.

Zhang YD, Ma J, Ma LG, Zhang XG. 2012. Two new species of Taeniolina from southern China.
Mycological Progress 11: 71-74. http://dx.doi.org/10.1007/s11557-010-0729-7

MY COTAXON

http://dx.doi.org/10.5248/130.47
Volume 130, pp. 47-56 January-March 2015

Fistulina subhepatica sp. nov. from China
inferred from morphological and sequence analyses

Jz Sonc, MEI-LING HAN, & BAo-Kai Cur

Institute of Microbiology, Beijing Forestry University, Beijing 100083, China
Beijing Key Laboratory for Forest Pest Control, Beijing Forestry University, Beijing 100083, China

* CORRESPONDENCE TO: cuibaokai@yahoo.com

ABSTRACT — Fistulina subhepatica sp. nov. is described from Yunnan Province, southwestern
China. The main characters of F subhepatica are its pileate basidiomata with rose to reddish
brown pileal surface and white pore surface, individual and easily separable pores (6-9
per mm), a monomitic hyphal structure with clamped generative hyphae, and ellipsoid
basidiospores (4-6 x 3-4 um). Phylogenetic analysis inferred from the combined ITS
(internal transcribed spacer region) and nLSU (the nuclear large subunit ribosomal RNA)
dataset support F. subhepatica as a distinct new species in Fistulina.

Key worps — Agaricales, Agaricomycetes, Fistulinaceae, phylogeny, rDNA, taxonomy

Introduction

Fistulina Bull. was established by Bulliard (1790: t.464) and typified by
F. buglossoides Bull. [= E hepatica (Schaeff.) With.]. It is a small but
cosmopolitan genus and causes a brown rot of hardwoods. Morphologically,
the genus is characterized by an annual growth habit, reddish to brown pileus
surface, separate but closely packed tubes, a monomitic hyphal system, and
ovoid basidiospores (Gilbertson & Ryvarden 1986, Nunez & Ryvarden 2001,
Ryvarden & Gilbertson 1993).

Previous multi-gene phylogenetic studies showed that Fistulina fell into the
euagarics clade and was closely related to Schizophyllum Fr., Auriculariopsis
Maire, and Porodisculus Murrill (Binder et al. 2005, Bodensteiner et al. 2004).

Taxonomic and phylogenetic studies focusing on brown-rot fungi in China
have been carried out recently, and some new species have been described (Cui
2013, Cui & Dai 2013, Cui & Li 2012, Cui et al. 2011, Li & Cui 2013a, Li et al.
2013, Shen et al. 2014, Zhou & Dai 2012, Zhou & Wei 2012). As a continuation
of these surveys, one undescribed species of Fistulina was discovered based on

48 ... Song, Han, & Cui

morphological characters and phylogenetic analysis of the ITS and nLSU rRNA
gene regions.

TABLE 1. List of species, vouchers, and GenBank accession numbers for sequences
used in phylogenetic analysis. New sequences are indicated by bold font.

GENBANK ACCESSION NO.

SPECIES VOUCHER nLsU ITS
Boletus satanas Lenz Bs 2 AF336242 DQ534567
TDB 1000c DQ533973
Coniophora olivacea (Fr.) P. Karst. 402 AF098376
Favolaschia andina Singer KG 0025 HM246679 HM246678
F. calocera R. Heim PDD 70689 AY572006
PDD 71528 AY572007
SR.KEN. 346 AF261417
E cf. calocera JM 98/372 AF261419
EF, peziziformis (Berk. & M.A. Curtis) Kuntze PDD 67440 AY572008
Favolaschia sp. KG 0027 HM246682 HM246681
Fistulina antarctica REG 550 AY571002
CBS 701.85 AY293181 DQ486702
CIEFAP 115 AY571003
EF. hepatica DSH 93-183 AY293182
REG 593 AY571004 AY571038
EF. pallida CBS 508.63 AY571005 AY571039
E. subhepatica Dai 11714 KJ925057
Dai 12416 KJ925053 KJ925058
Cui 11130 KJ925054 KJ925059
Dai 13216 KJ925055 KJ925060
Dai 13244 (HT) KJ925056 KJ925061
Jaapia argillacea CBS 252.74 GU187581 GU187524
Mycena epipterygia (Scop.) Gray GLM 45969 AY207249
M. galericulata (Scop.) Gray GLM 45970 AY207251
TENN 7495 AF261412
RV 87/14.01 AF042636
M. pura (Pers.) P. Kumm. CBH 371 FN394630 KF913023
IS 10/11/2000 FN394634 FN394611
CBH 367 FN394631 KF913022
CBH 358 FN394629 FN394608
M. rosea Gramberg CBH 097 FN394628 FN394556
TL 12393 FN394641 FN394555
UP 2 FN394632 FN394550
M. sanguinolenta (Alb. & Schwein.) P. Kumm. 430360 AB512401 AB512311
Panellus edulis Y.C. Dai et al. HMJAU 7066 GQ219739 GQ219731
HMJAU 7214 GQ219738 GQ219730

Fistulina subhepatica sp. nov. (China) ... 49

Paxillus involutus (Batsch) Fr. UBC F16285 EU486436

Poromycena manipularis (Berk.) R. Heim JM 98/217 AF261423

Porodisculus pendulus HHB-15376-sp AY293204
DAOM 198417 AF261593

Poromycena sp. JM 98/128 AF261429

Suillus cavipes (Opat.) A.H. Sm. & Thiers TDB-646 AF071535

Schizophyllum amplum (Lév.) Nakasone FCUG 1803 AF141873

S. commune Fr. NBRC 4928 AB733339 AB733163
NBRC 6502 AB733340 AB733164
NBRC 30496 AB733341 AB733165
NBRC 30749 AB733342 AB733166

S. fasciatum Pat. CBS 267.60 AF261589 L43385

S. umbrinum Berk. FL 02.1 AF261590 AF249391

HT = holotype

Materials & methods

Morphological studies

The studied specimens are deposited at the herbarium of the Institute of
Microbiology, Beijing Forestry University (BJFC). Microscopic examinations followed
Dai et al. (2010). Sections were studied at magnifications up to 1000x using a Nikon
E 80i microscope and phase contrast illumination (Nikon, Tokyo, Japan). Drawings
were made with the aid of a drawing tube. Microscopic features, measurements, and
drawings were made from slide preparations stained with cotton blue and Melzer’s
reagent. Basidiospores were measured from sections cut from the tubes. In presenting
the variation in the basidiospore size, 5% of measurements that were excluded from
each end of the range are given in parentheses. In the text, the following abbreviations
were used: IKI = Melzer’s reagent, IKI- = negative in Melzer’s reagent, KOH = 5%
potassium hydroxide, CB = cotton blue, CB- = acyanophilous, CB+ = cyanophilous,
L = mean spore length (arithmetic average of all spores), W = mean spore width
(arithmetic average of all spores), Q = variation in the L/W ratios among the specimens
studied, n = number of spores measured from given number of specimens. Special color
terms follow Petersen (1996).

Molecular procedures and phylogenetic analysis

The Phire Plant Direct PCR Kit (Finnzymes, Vantaa, Finland) was used to obtain PCR
products from dried specimens, according to the manufacturer's instructions. A small
piece of dried fungal specimen was lysed in 30 ul dilution buffer for DNA extraction.
After incubating for 3 min at room temperature, 0.75 ul of the supernatant was used
as template for a 30 ul PCR reaction. The internal transcribed spacer (ITS) regions
were amplified with the primers ITS4 and ITS5 (White et al. 1990), and the nuclear
large subunit (nLSU) ribosomal RNA gene regions with the primers LROR and LR7
(http://www. biology.duke.edu/fungi/mycolab/primers.htm). The PCR procedure for
ITS was as follows: initial denaturation at 95°C for 3 min, followed by 34 cycles at 94°C
for 40 s, 54°C for 45 s, and 72°C for 1 min, and a final extension of 72°C for 10 min. The

50 ... Song, Han, & Cui

PCR procedure for nLSU was as follows: initial denaturation at 94°C for 1 min, followed
by 34 cycles at 94°C for 30 s, 50°C for 1 min and 72°C for 1.5 min, and a final extension
of 72°C for 10 min. The PCR products were purified and directly sequenced in Beijing
Genomics Institute, China, with the same primers. All newly generated sequences were
submitted to GenBank (TABLE 1).

Sequences generated for this study were aligned with additional sequences
downloaded from GenBank (TaBLE 1) using BioEdit (Hall 1999) and ClustalX
(Thompson et al. 1997). Prior to phylogenetic analysis, ambiguous positions at the
start and the end were deleted and gaps were manually adjusted to optimize alignment.
Sequence alignment was deposited at TreeBASE (http://purl.org/phylo/treebase;
submission ID 15918).

Phylogenetic analysis was done as in Li & Cui (2013b). Maximum parsimony
analysis and Bayesian inference (BI) was applied to the combined ITS and nLSU dataset.
Jaapia argillacea Bres. was used as outgroup to root trees (Bodensteiner et al. 2004). The
maximum parsimony tree was constructed in PAUP* version 4.0b10 (Swofford 2002).
All characters were equally weighted and gaps were treated as missing data. Trees were
inferred using the heuristic search option with TBR branch swapping and 1000 random
sequence additions. Max-trees were set to 5000, branches of zero length were collapsed
and all parsimonious trees were saved. Clade robustness was assessed using a bootstrap
(BT) analysis with 1000 replicates (Felsenstein 1985). Descriptive tree statistics tree
length (TL), consistency index (CI), retention index (RI), rescaled consistency index
(RC), and homoplasy index (HI) were calculated for each maximum parsimonious tree
(MPT) generated.

MrMODELTEST2.3 (Nylander 2004, Posada & Crandall 1998) was used to
determine the best-fit evolution model for BI. BI was calculated with MrBayes3.1.2
(Ronquist & Huelsenbeck 2003) according to the calculated model. Four Markov chains
were run for 2 runs from random starting trees for 1.5 million generations, and trees
were sampled every 100 generations. The first one-fourth generations were discarded as
burn-in. A majority rule consensus tree of all remaining trees was calculated. Branches
that received bootstrap support for maximum parsimony (MP) 275% and Bayesian
posterior probabilities (BPP) 20.95 were considered as significantly supported.

Results

The combined ITS and nLSU dataset include sequences from 49 fungal
collections. The dataset has an aligned length of 1388 characters including
gaps, of which 812 characters are constant, 135 are parsimony-uninformative,
and 441 are parsimony-informative. Maximum parsimony analysis yielded
9 equally parsimonious trees (Length = 1373, CI = 0.650, RI = 0.867, RC = 0.564,
HI = 0.350), and a strict consensus tree of these trees is shown in Fic. 1. The
best model for the combined ITS and nLSU partition is a GTR+I+G model. BI
produced a topology similar to that from MP analysis with an average standard
deviation of split frequencies = 0.007406.

In the phylogenetic tree (Fic. 1), the unknown Chinese Fistulina collections
grouped in a well-supported lineage (MP = 88%, BPP = 1.00), which had a

Fistulina subhepatica sp. nov. (China) ... 51

Favolaschia calocera SR.KEN. 346
a Soins Favolaschia calocera PDD 70689
97/1.00 deleiay Favolaschia calocera PDD 71528
-/0.98| © Favolaschia cf. calocera JM 98/372
IN Favolaschia sp. KG 0027
78/1.00] Favolaschia andina KG 0025
Favolaschia peziziformis PDD 67440
Poromycena manipularis JM 98/217
59/- ||; Mycena galericulata TENN 7495
Mycena galericulata GLM 45970
Mycena epipterygia GLM 45969
Mycena rosea CBH 097
98/0.97 ae Mycena rosea UP 2
Mycena rosea TL 12393
LK) Mycena pura CBH 371
100/1.00 Mycena pura CBH 367
Mycena pura CBH 358
Mycena pura I§ 10/11/2000
89/1.00 Mycena sanguinolenta 430360
Poromycena sp. JM 98/128
Mycena galericulata RV 87/14.01
100/1.00 Panellus edulis HMJAU 7066
Panellus edulis HMJAU 7214
Fistulina subhepatica Dai 11714
Fistulina subhepatica Dai 13244

100/-

99/1.00
88/1.00) Fistulina subhepatica Dai 13216
87/0.99 100/1.00 Fistulina subhepatica Cut 11130
Fistulina subhepatica Dai 12416 Fi stu lin a
100/1.00 99/199 Fistulina hepatica REG 593
Fistulina hepatica DSH 93-183
Fistulina antarctica REG 550
100/1.00 Fistulina antarctica CBS 701.85
98/0.99 100/1.00 64/- Fistulina antarctica CIEFAP 115
Porodisculus pendulus HHB-15376-sp
Porodisculus pendulus DAOM 198417
93/0.99 L_. Fistulina pallida CBS 508.63
Schizophyllum commune NBRC 30749
90/- Schizophyllum commune NBRC 30496
58/- 89/0.99 pelzepioiiuin commune NBRC 4928
Schizophyllum commune NBRC 6502
Schizophyllum fasciatum CBS 267.60
100/1.00

Schizophyllumumbrinum FL 02.1
79/- © Schizophyllumamplum FCUG 1803
66/0.99 — Boletus satanas Bs 2
91/0.98]L Boletus satanas TDB 1000c
89/1.00 Paxillus involutus UBC F16285
Suillus cavipes TDB-646

Coniophora olivacea 402

Jaapia argillacea CBS 252.74

100/-

Figure 1. The phylogenetic position of Fistulina subhepatica based on ITS+nLSU sequence data.
Topology is from maximum parsimony analysis. Bootstrap values >50% and Bayesian posterior
probabilities 20.95 are indicated at the nodes.

sister relationship with E hepatica [= F buglossoides, the generic type]. The new
species is described on the next page.

52... Song, Hany-& Cui

FiGurRE 2. Fistulina subhepatica (holotype), basidiomata in situ. Scale bar = 2 cm.

Taxonomy

Fistulina subhepatica B.K. Cui & J. Song, sp. nov. FIGs. 2-3
MycoBank MB 809111
Differs from Fistulina hepatica by its thin- to slightly thick-walled tramal generative

hyphae with clamp connections and larger basidiospores.

Type: China, Yunnan Province, Jingdong County, Ailaoshan Nature Reserve, on
dead tree of Lithocarpus (Fagaceae), 12.V1I.2013, Dai 13244 (holotype, BJFC 014732;
GenBank KJ925056, KJ925061).

EryMoLoecy: subhepatica (Lat.): refers to the morphological similarity to Fistulina

hepatica.
FruiTBopy — Basidiomata annual, pileate, fleshy, and readily exuding a
reddish blood-like sap when squeezed or bruised when fresh, leathery when
dry. Pileus sessile to laterally substipitate, dimidiate to reniform or subcircular,
projecting up to 25 cm, 30 cm wide, and 6 cm thick at base. Pileal surface rose
to reddish brown when fresh, becoming fuscous to black upon drying, faintly
radially furrowed when fresh; margin acute, concolorous. Pore surface white
when fresh, turning darker when bruised and becoming cinnamon-buff to
reddish brown when dry; pores 6-9 per mm, consisting of individual, crowed
but easily separable tubes. Context clay-buff to black, leathery when dry, up
to 5 cm thick. Tubes concolorous with pore surface, leathery, up to 1 cm long.

Fistulina subhepatica sp. nov. (China) ... 53

iabaateiae aes
10 pm 10 ym

SSS
10 ym

FiGure 3. Fistulina subhepatica (holotype). a: Basidiospores. b: Basidia and basidioles.
c: Cystidioles. d: Cystidial elements from dissepiment edges. e: Hyphae from trama.
f: Hyphae from context. Scale bars = 10 um.

54 ... Song, Han, & Cui

HYPHAL STRUCTURE — Hyphal system monomitic; generative hyphae with
clamp connections, IKI-, CB-, more or less dissolving in KOH.

CONTEXT — Generative hyphae hyaline, thin-walled, rarely branched,
interwoven, 5-12 um in diam, but with inflated portions up to 18 um,
gloeoplerous hyphae present.

TuBES — Generative hyphae hyaline, thin- to slightly thick-walled,
rarely branched, parallel along the tubes and densely in sections from dried
specimens, 3.5-8 um in diam. Basidia clavate with four sterigmata and a basal
clamp connection, 15-32 x 5-7 um; basidioles in shape similar to basidia, but
slightly smaller. Cystidial elements present in the dissepimental edges, hyaline,
thin-walled, 75-90 x 5-7 um.

Spores — Basidiospores ellipsoid, hyaline, thick-walled, smooth,
IKI-, CB+, 4-6 x 3-4(-4.5) um, L = 4.82 um, W = 3.37 um, Q = 1.36-1.52
(n = 150/5).

ADDITIONAL SPECIMENS EXAMINED — CHINA. YUNNAN PROVINCE, LINCANG,
Xiaodaohe Forest Farm, on dead tree of Castanopsis (Fagaceae), 10.V1I.2013, Dai 13216
(BJFC 014706; GenBank KJ925055, KJ925060); NANHUA County, Dazhongshan Nature
Reserve, on fallen angiosperm trunk, 15.VII.2013, Cui 11130 (BJFC 015245; GenBank
KJ925054, KJ925059); CHuxIoNG, Zixishan Forest Park, on dead tree of Lithocarpus,

28.VUI.2010, Dai 11714 (BJFC 008827; GenBank KJ925057); 11.VI.2011, Dai 12416
(BJFC 010696; GenBank KJ925053, KJ925058).

OTHER SPECIMEN EXAMINED — Fistulina hepatica. FINLAND. VANTAA, Tammosto
Nature Reserve, on dead tree of Quercus (Fagaceae), 8.1X.2012, Dai 12874 (BJFC
013153).

Discussion

The previous reports of Fistulina hepatica in China (Dai 2012, Dai et al.
2011, Wang et al. 2011, Yuan & Dai 2008) were from identifications based only
on morphological characters. According to the current phylogenetic analysis
and more careful morphological examinations, these Chinese samples are
newly described as F. subhepatica. Macroscopically, FE hepatica is quite similar
to FE. subhepatica in its pinkish brown to more reddish or purplish brown pileal
surface, white pore surface when fresh, and individual pores; microscopically,
however, F. hepatica differs from F. subhepatica by having smaller basidiospores,
simple septate and thin-walled trama generative hyphae (Gilbertson &
Ryvarden 1986, Nunez & Ryvarden 2001, Ryvarden & Gilbertson 1993).
Moreover, F. hepatica forms a distinct lineage separate from F. subhepatica
(Fic. 1). In addition, E hepatica is widely distributed in temperate Europe and
grows mainly on Quercus and Castanea, whereas F. subhepatica is found in
subtropical China, occurring mostly on Castanopsis and Lithocarpus. We did
not find FE hepatica on Quercus in northern China, although the big and old
oak trees are suitable for the fungus.

Fistulina subhepatica sp. nov. (China) ... 55

Fistulina antarctica Speg., closely related to F. subhepatica in the phylogenetic
analysis, differs morphologically from FE subhepatica by its longer basidiospores
(5-7 x 3-4 um, Spegazzini 1887).

Fistulina pallida Berk. & Ravenel resembles F. subhepatica in its brownish
pore surface and individual pores; however, it produces smaller basidiomata and
wood-brown pileus surface (Gilbertson & Ryvarden 1986). Phylogenetically,
E pallida falls outside the “core Fistulina clade” and groups with Porodisculus
pendulus (Fr.) Murrill.

Acknowledgments

The authors are grateful to Prof. Yu-Cheng Dai (BJFC, China) for collecting
specimens and improving the text. We express our gratitude to Drs. Li-Wei Zhou
(China) and Tatiana B. Gibertoni (Brazil), who reviewed the manuscript. The research
was financed by Beijing Higher Education Young Elite Teacher Project (YETP0774) and
the National Natural Science Foundation of China (Project No. 31170018).

Literature cited

Binder M, Hibbett DS, Larsson KH, Larsson E, Langer E, Langer G. 2005. The phylogenetic
distribution of resupinate forms across the major clades of mushroom-forming fungi
(Homobasidiomycetes). Systematics and Biodiversity 2: 113-157.
http://dx.doi.org/10.1017/s1477200005001623

Bodensteiner P, Binder M, Moncalvo JM, Agerer R, Hibbett DS. 2004. Phylogenetic relationships
of cyphelloid Homobasidiomycetes. Molecular Phylogenetics and Evolution 2: 501-515.
http://dx.doi.org/10.1016/j.ympev.2004.06.007

Bulliard JBE. 1790. Herbier de la France 10: t.433-480.

Cui BK. 2013. Antrodia tropica sp. nov. from southern China inferred from morphological
characters and molecular data. Mycological Progress 12: 223-230.
http://dx.doi.org/10.1007/s11557-012-0829-7

Cui BK, Dai YC. 2013. Molecular phylogeny and morphology reveal a new species of
Amyloporia (Basidiomycota) from China. Antonie van Leeuwenhoek 104: 817-827.
http://dx.doi.org/10.1007/s10482-013-9994-1

Cui BK, Li HJ. 2012. A new species of Postia (Basidiomycota) from Northeast China. Mycotaxon
120: 231-237. http://dx.doi.org/10.5248/120.231

Cui BK, Li HJ, Dai YC. 2011. Wood-rotting fungi in eastern China 6. Two new species of
Antrodia (Basidiomycota) from Mt. Huangshan, Anhui Province. Mycotaxon 116: 13-20.
http://dx.doi.org/10.5248/116.13

Dai YC. 2012. Polypore diversity in China with an annotated checklist of Chinese polypores.
Mycoscience 53: 49-80. http://dx.doi.org/10.1007/s10267-011-0134-3

Dai YC, Cui BK, Liu XY. 2010. Bondarzewia podocarpi, a new and remarkable polypore from
tropical China. Mycologia 102: 881-886. http://dx.doi.org/10.3852/09-050

Dai YC, Cui BK, Yuan HS, He SH, Wei YL, Qin WM, Zhou LW, Li HJ. 2011. Wood-inhabiting fungi
in southern China 4. Polypores from Hainan Province. Annales Botanici Fennici 48: 219-231.
http://dx.doi.org/10.5735/085.048.0302

Felsenstein J. 1985. Confidence intervals on phylogenetics: an approach using bootstrap. Evolution
39:783-791,

Gilbertson RL, Ryvarden L. 1986. North American polypores 1. Fungiflora, Oslo.

56 ... Song, Han, & Cui

Hall TA. 1999. Bioedit: a user-friendly biological sequence alignment editor and analysis program
for Windows 95/98/NT. Nucleic Acids Symposium Series 41: 95-98.

Li HJ, Cui BK. 2013a. Two new Daedalea species (Polyporales, Basidiomycota) from South China.
Mycoscience 54: 62-68. http://dx.doi.org/10.1016/j.myc.2012.07.005

Li HJ, Cui BK. 2013b. Taxonomy and phylogeny of the genus Megasporoporia and its related genera.
Mycologia 105: 368-383. http://dx.doi.org/10.3852/12-114

Li HJ, Han ML, Cui BK. 2013. Two new Fomitopsis species from southern China based
on morphological and molecular characters. Mycological Progress 12: 709-718.
http://dx.doi.org/10.1007/s11557-012-0882-2

Nunez M, Ryvarden L. 2001. East Asia polypores 2. Polyporaceae s. lato. Synopsis Fungorum 14:
341-342.

Nylander JAA. 2004. MrModeltest v2. Program distributed by the author. Evolutionary Biology
Centre, Uppsala University.

Petersen JH. 1996. Farvekort. The Danish Mycological Society's colour-chart. Foreningen til
Svampekundskabens Fremme, Greve. 6 p.

Posada D, Crandall KA. 1998. Modeltest: testing the model of DNA substitution. Bioinformatics
14: 817-818. http://dx.doi.org/10.1093/bioinformatics/14.9.817

Ronquist F, Huelsenbeck JP. 2003. MRBAYES 3: bayesian phylogenetic inference under mixed
models. Bioinformatics 19: 1572-1574.

Ryvarden L, Gilbertson RL. 1993. European polypores 1. Synopsis Fungorum 6: 372-374.

Shen LL, Cui BK, Dai YC. 2014. A new species of Postia (Polyporales, Basidiomycota) from
China based on morphological and molecular evidence. Phytotaxa 162: 147-156.
http://dx.doi.org/10.11646/3529

Spegazzini C. 1887. Fungi Patagonici. Boletin de la Academia Nacional de Ciencias en Cordoba 11:
5-64. http://dx.doi.org/10.5962/bhl.title.4071

Swofford DL. 2002. PAUP*: phylogenetic analysis using parsimony (*and other methods). version
4.0b10. Sinauer Associates, Sunderland, Massachusetts.

Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG. 1997. The Clustal_X windows
interface: Flexible strategies for multiple sequence alignment aided by quality analysis tools.
Nucleic Acids Research 25: 4876-4882. http://dx.doi.org/10.1093/nar/25.24.4876

Wang B, Cui BK, Li HJ, Du P, Jia BS. 2011. Wood-rotting fungi in eastern China 5.
Polypore diversity in Jiangxi Province. Annales Botanici Fennici 48: 237-246.
http://dx.doi.org/10.5735/085.048.0304

White TJ, Bruns T, Lee S, Taylor J. 1990. Amplification and direct sequencing of fungal ribosomal
RNA genes for phylogenetics. 315-322, in: MA Innis et al. (eds). PCR Protocols: a guide to
methods and applications. Academic, San Diego.

Yuan HS, Dai YC. 2008. Polypores from northern and central Yunnan Province, Southwestern
China. Sydowia 60: 147-159.

Zhou LW, Dai YC. 2012. Progress report on the study of wood-decaying fungi in China. Chinese
Science Bulletin 57: 4328-4335. http://dx.doi.org/10.1007/s11434-012-5457-8

Zhou LW, Wei YL. 2012. Changbai wood-rotting fungi 16. A new species of Fomitopsis
(Fomitopsidaceae). Mycological Progress 11: 435-441.
http://dx.doi.org/10.1007/s11557-011-0758-x

MYCOTAXON

http://dx.doi.org/10.5248/130.57
Volume 130, pp. 57-59 January-March 2015

Antherospora sukhomlyniae, a new species of smut fungi on
Hyacinthella in Crimea (Ukraine)

KYRYLO G. SAVCHENKO*

Department of Plant Pathology, Washington State University,
Pullman, WA 99164, United States

* CORRESPONDENCE TO: kyryll.savchenko@wsu.edu

AsBstTRACT — Antherospora sukhomlyniae, a new species of smut fungus parasitic on
Hyacinthella sp. in Crimea (Ukraine), is described and illustrated.

Key worps — anther smut, biodiversity, mycobiota, Ustilaginomycetes

Introduction

During a collecting trip for smut fungi in Crimea (Ukraine), carried out
by the author in 2009, several specimens of smut fungi were collected. Based
on the study of the specimen from anthers of Hyacinthella sp. (Asparagaceae),
I propose here a new smut fungus, Antherospora sukhomlyniae, that was
collected in Karadag reserve in the southeastern part of the Crimean peninsula.

Materials & methods

Sorus and spore characteristics were studied using dried herbarium specimens.
Spores were dispersed in a droplet of lactic acid on a microscope slide, covered with
a cover glass, gently heated to boiling point to rehydrate the spores, cooled, and then
examined using a Carl Zeiss Axiostar light microscope (LM) at 1000x magnification.
For scanning electron microscopy (SEM), spores were attached to specimen holders
by double-sided adhesive tape and coated with gold. The surface structure of spores
was observed at 15 kV and photographed with a scanning electron microscope JEOL
JSM-6700EF.

The holotype specimen is conserved in the Herbarium, Department of Biodiversity
and Biotechnology of Fungi, Institute of Evolution, University of Haifa, Israel (HAI).

Taxonomy

Antherospora sukhomlyniae K.G. Savchenko, sp. nov. FIG. 1
MycoBAnk 808616

58 ... Savchenko

FiGurE 1. Antherospora sukhomlyniae (holotype) in flowers of Hyacinthella sp.: spores. a = LM;
b-d = SEM. Bars: a, b= 10 um, c = 2 um, d= 1 um.

Differs from all other Antherospora species by its host specialization on Hyacinthella and
from A. albucae and A. urgineae by its smaller spores, from A. eucomis by its irregularly
shaped spores, and from A. tourneuxii by its thicker spore walls.

Type: Ukraine, Autonomous Republic of Crimea, Karadag reserve, in flowers of
Hyacinthella sp., 5.V.2009, leg. K.G. Savchenko (Holotype HAI 6521).

EryMo_oey: named in honor of the Ukrainian mycologist Maryna M. Sukhomlyn.

Parasitic on Hyacinthella. Sori in all inner floral organs of deformed and
swollen flowers, producing blackish brown, powdery mass of spores, for a long
time enclosed by the outermost floral envelopes. Infection systemic, with all
flowers of the plant infected. Spores ovoid, ellipsoidal, subglobose to irregular,
(6.5-)7-10 x 7.5-13(-14) um, yellowish brown. Spore wall even, ca. 0.8 um
thick, finely densely punctate-verrucose, with finely wavy spore profile.

Antherospora sukhomlyniae sp. nov. (Ukraine) ... 59

DISTRIBUTION - Known only from the type locality in Crimea, on
Hyacinthella sp.

COMMENTS -— Previous studies have demonstrated that while all species of
Antherospora attack hosts in the Asparagaceae, they display a high level of
specificity, with each species restricted to host plants from a single genus, or in
some cases, from a single species (Bauer et al. 2008, Piatek et al. 2011, 2013).
As no Antherospora species has been reported on Hyacinthella, 1 propose a new
species here.

Antherospora sukhomlyniae belongs to the group of Antherospora species
with sori destroying all inner floral organs of infected flowers (Vanky 2012),
which includes A. albucae (Syd. & P. Syd.) R. Bauer et al., A. eucomis Vanky, A.
tourneuxii (A.A. Fisch. Waldh.) R. Bauer et al., and A. urgineae (Maire) R. Bauer
et al. However, these species can be distinguished from A. sukhomlyniae: A.
albucae by its larger spores (7-14.5 x 9.5-22.5 um; Vanky 2012); A. urgineae by
its larger spores (7-12 x 9.5-17.5 um; Vanky 2012); A. eucomis by its regularly
shaped spores with thinner spore walls (0.5 um; Vanky 2009); and A. tourneuxii
by its thinner spore walls (0.5 um; Vanky 2012).

Acknowledgments

The author thanks Lori M. Carris and Vasyl P. Heluta for peer-reviewing the
manuscript, Shaun Pennycook for useful corrections, and Vitalii Sapsai for help with
the SEM microscopy.

Literature cited

Bauer R, Lutz M, Begerow D, Piatek M, Vanky K, Bacigalova K, Oberwinkler F. 2008. Anther smut
fungi on monocots. Mycological Research 112: 1297-1306.
http://dx.doi.org/10.1016/j.mycres.2008.06.002

Piatek M., Lutz M., Chater A.O. 2013. Cryptic diversity of the Antherospora vaillantii complex on
the Muscari species. IMA Fungus 4: 5-19.

Piatek M., Lutz M., Smith P.A., Chater A.O. 2011. A new species of Antherospora supports the
systematic placement of its host plant. IMA Fungus 2: 135-142.

Vanky K. 2009. Taxonomic studies on Ustilaginomycetes - 29. Mycotaxon 110: 289-324.
http://dx.doi.org/10.5248/110.289

Vanky K. 2012. Smut fungi of the world. APS Press, USA.

MY COTAXON

http://dx.doi.org/10.5248/130.61
Volume 130, pp. 61-68 January-March 2015

Tuber xanthomonosporum,
a new Paradoxa-like species from China

YUAN QING", SHU-HONG LI’, CHENG-YI Liv?, LIN Li’, MEI YANG},
XIAO-LEI ZHANG’, XIAO-LIN L14, LIN- YONG ZHENG?* 4, & YUN WANG®®

' Key Laboratory of Bio-resources and Eco-environment, Ministry of Education, School of Life
Science, Sichuan University, 24 (South part) First Ring Road, Chengdu, Sichuan, China

? Biotechnology & Germplasm Resources Institute, Yunnan Academy of Agricultural Sciences,
9 Xueyun Road, Kunming, Yunnan, China

Sichuan Panzhihua Academia of Agriculture and Forestry,
1719 Panzhihua Road, Panzhihua, Sichuan, China

‘Sichuan Academy of Agricultural Sciences, 20 Jingjusi Road, Chengdu, Sichuan, China

‘Department of Light Chemical Engineering, Xichang College,
Mapingba Road, Xichang, Sichuan, China

°Yunnan Institute for Tropical Crop Research, 99 Xuanwei Road, Jinghong, Yunnan, China

* CORRESPONDENCE TO: “zly6559@126.com "wangynz@yahoo.com

ABSTRACT — A new species, Tuber xanthomonosporum, is described based on specimens
collected under Pinus yunnanensis in Panzhihua, Sichuan Province, China. Tuber
xanthomonosporum invariably has only one spore per ascus. It can be distinguished from
the three other Chinese Paradoxa-like species — T: gigantosporum, T: sinomonosporum, and
T. glabrum — by its whitish to yellow-brownish gleba, two-layered peridium, and spiky
cystidia. Molecular analysis also supports T: xanthomonosporum as a unique species.

KEY worps — taxonomy, Pezizales, Yunnan

Introduction

The genus Paradoxa was erected by Mattirolo in 1935 in the Tuberaceae
to accommodate Paradoxa monospora Mattir., a species which, invariably,
has one-spored asci (Montecchi & Sarasini 2000; Leessoe & Hansen 2007).
For the following 50 years the genus remained monotypic until Wang & Hu
(2008) transferred Tuber gigantosporum into Paradoxa. Wang & Li (1991)
originally described T’ gigantosporum based on a single specimen collected
from Huidong County, Sichuan, China, in 1988. The species was characterized
by its extremely large elliptical ascospores and one-spored asci, a feature that
was eventually recognized as being important when a second T: gigantosporum

62 ... Qing & al.

collection was made in Yongren County, Yunnan, China, in 2002. Since then
three other Paradoxa-like species have been reported: Tuber sp. 1 (Kinoshita
et al. 2011), Paradoxa sinensis (Fan et al. 2012), and Tuber glabrum (Fan et
al. 2014). Recent molecular phylogenetic analysis of truffle species supported
Paradoxa within Tuber (Kinoshita et al. 2011, Fan et al. 2013), and subsequently
Fan et al. (2014) transferred P sinensis into Tuber with the replacement name

T. sinomonosporum.

In 2012 during a survey in Panzhihua in the southwestern corner of Sichuan,
China, we found truffles that invariably had one-spored asci. Here we describe
this Paradoxa-like truffle as a new species of Tuber and briefly review the

molecular relationships of truffle species with single-spored asci.

TABLE 1: Origin of the fungal ITS sequences. (New sequences are indicated

in bold font.)

TAXON

Choiromyces meandriformis

Gymnohydnotrya
australiana

Tuber sp. 1
Tuber sp. 2
T. borchii

T. canaliculatum

T. excavatum

T. foetidum

T. fulgens

T. glabrum

T. indicum

T. latisporum

VOUCHER OR CODE

isolate RH691
isolate GB285

isolate OSC130601

isolate K201
isolate K215

clone Bo6

strain Z43
Unknown
JT12670

JT23942

TE-M

isolate zb3281_215
TE-WD

isolate B-2452
isolate B-2489
isolate y466

isolate zb3386_202
isolate M2435

strain T39
strain T47
strain T49
HKAS 44315
HKAS 42380

GEOGRAPHICAL ORIGIN

USA
USA

USA

Japan
Japan
Italy
Italy
France
USA
USA
Poland
Germany
Poland
Hungary
Hungary
Germany
Germany
USA
China
China
China
China
China
China
China

GENBANK NO.

HM485330
HM485331

JQ925629

AB553344
AB553356
AF106890
AF250291
AF132505
GQ221455
GQ221456
KC330228
HM152020
KC330227
AJ557543
AJ557544
HM152014
HM152024
HM485358
KFO002731
KF002732
JQ638997
JQ639005
JQ639007
DQ898183
DQ898184

Tuber xanthomonosporum sp. nov. (China) ... 63

T. liui HKAS 48269 China DQ898182
T. macrosporum isolate JT 19458 USA HM485372
isolate $7510 Germany JF926121
T. maculatum clone Macl Italy AF106889
1967 New Zealand EU753269
T. melanosporum strainl015 Israel AF167096
LHB-mic-Tmel-Qc-09-1 Spain GU810153
LHB-mic-Tmel-Qr-09-1 Spain GU810152
T. panzhihuanense strain DXJ260 China JQ978644
strain DXJ276 China JQ978650
T. pseudoexcavatum isolate CJ408 USA HM485381
strain E China JQ638958
strain Q China JQ638982
strain T48 China JQ639006
T. rufum 1798 Italy EF362473
1447 Italy EF362477
T. sinomonosporum BJTCFAN150 China KF002729
T. umbilicatum HKAS48267 China GU979032
HKAS44316 China GU979031
isolate T104 China FJ797879
isolate T117 China FJ797880
T. xanthomonosporum YAAS 13185 China KJ162154
YAAS L3186 China KJ162155
YAAS L3187 China KJ162156
T. zhongdianense HKAS 45388B China DQ898186

wang0299 China DQ898187

Materials & methods

Morphological studies

Specimens of a truffle with asci containing a single spore were collected from
Panzhihua, Sichuan, China, in 2012. Specimens are deposited in the Herbarium of
Biotechnology and Germplasm Resources Institute, Yunnan Academy of Agricultural
Sciences, Kunming, Yunnan, China (YAAS).

The samples were examined at the Biotechnology and Germplasm Resources
Institute, Yunnan Academy of Agricultural Sciences, Kunming, China and Sichuan
Panzhihua Academy of Agriculture and Forestry, Panzhihua, Sichuan, China.
Macroscopic characters were described from fresh specimens and microscopic features
described from dried specimens following the methods of Yang & Zhang (2003).
Photographs were made under a Nikon E400 microscope. For scanning electron
microscopy (SEM), spores were scraped from the dried gleba onto double-sided tape
and mounted directly on a SEM stub. They were then coated with gold-palladium and
examined and photographed with a JEOL, JMS-5600LV SEM.

64 ... Qing & al.

DNA amplification & sequence analysis

DNA was extracted using CTAB (Doyle 1987) modified by adding 200 wL 5M
potassium acetate after 4 x CTAB treatments. The primers ITSIF (Gardes & Bruns
1993) and ITS4 (White et al. 1990) were used to amplify the ITS-rDNA region. PCR
reaction solution and cycling parameters used by Chen & Liu (2007) were adopted.
The amplification products were electrophoresed on a 1% agarose gel and purified with
Sangons purification kit. Sequencing was performed with a BigDye® Terminator v3.1
Cycle Sequencing Kit on an ABI 3730XL automatic sequencer.

In order to determine whether the specimens represented a new species, we compared
our Tuber xanthomonosporum ITS-rDNA sequences with 48 ITS-rDNA sequences
downloaded from NCBI. An ITS-rDNA sequence of Gymnohydnotrya australiana B.C.
Zhang & Minter was also included as outgroup. Sequence alignment and phylogenetic
analysis protocols followed Chen & Liu (2007).

Results

Phylogenetic analysis

A total of 52 ITS sequences were included in a phylogenetic analysis
(TABLE 1, PLATE 2). Of the 712 total characters analyzed, 248 are constant,
51 parsimony-uninformative, and 413 parsimony-informative (L = 1979,
CI = 0.481, RI = 0.812, RC = 0.390). ITS sequence analysis revealed that the
three Paradoxa-like species (Tuber sp. 1, T: sinomonosporum, T. glabrum) form
a well-supported subclade (bootstrap support = 98%) within Clade I, which
also included T: macrosporum, T. canaliculatum, and Tuber sp. 2. In contrast,
T. xanthomonosporum does not cluster with the other Paradoxa-like species
but forms a new Subclade I in Clade II (with low bootstrap support) alongside
Subclade II (/melanosporum) and Subclade III (/rufum).

Taxonomy

Tuber xanthomonosporum Qing & Y. Wang, sp. nov. PLATE 1
MycoBAnk MB 807668

Differs from other Tuber species with single-spored asci by its whitish to yellow-
brownish gleba and two-layered peridium with spiky cystidia.

TyPE: China, Sichuan Province, Panzhihua, 26°29’N 102°01’E, under Pinus yunnanensis
Franch., 15 Nov. 2012: Y. Qing Pan1201 (Holotype, YAAS L3185; GenBank KJ162154).

EryMoLoecy: xanthomonosporum from Latin refers to the yellow brownish gleba and
single spored asci.

PiaTE 1. Tuber xanthomonosporum (YAAS L3185, holotype): 1, 2. Fresh ascomata showing
surface features and cut sections. 3. Cross section of peridium showing two layers of pseudo-
parenchymatous and interwoven tissue with some cystidia. 4. Cross section of outer layer of
peridium showing two kinds of cystidia: talland tapered (arrows), and short and obtuse (arrowhead).
5-7. Ascospores and asci (invariably with only one spore). 8. Ascospores showing ornamentations.
Scale bars: 1, 2 = 1 cm; 3, 8 = 20 um; 4, 7 = 50 um; 5 = 100 um; 6 = 40 um.

Sh 14 (20°

66 ... Qing & al.

Ascomata subglobose or irregular and much lobed with deep furrows, firm
and a little rubbery, whitish to yellow-brownish, up to 3.0 cm broad, rough
with hairs. Opor slightly garlicky when mature. PERIDIuM 160-200 um
thick, composed of two layers; the outer layer complex, 50-70 um thick,
pseudoparenchymatous, composed of irregular globose to rectangular cells,
19-32 x 13-15 um diam and thin-walled; the outmost cells giving rise at the
surface to cystidia of two types; one tall, thin, tapered, reaching 50-90 x 2-4 um
and sometimes with a small teat on the wall, hyaline; the other short, obtuse,
1-2-septate, 15-20 x 4-5 um; the inner layer 110-130 um thick, composed of
mainly interwoven, colourless hyphae of 2-4 um diam with some big cells of
2-4 um diam, that gradually merge into the gleba tissue. GLEBA solid, whitish
when young, becoming yellow-brownish at maturity, marbled with distinct,
white and meandering veins, merging at many points with the peridium. Asc1
60 x 70 um, globose to subglobose, sessile, slightly thick walled 2-3 um thick,
invariably 1-spored and randomly dispersed in glebal tissue. AscospoREs
globose, 41-43 x 43-46 um; spore walls 2.5 um thick, brown at maturity,
ornamented with a regular alveolate-reticulum, 2-3 um deep, constituted of
mostly hexagonal meshes 8-15 x (5-)7-13 um, 4-5 across the spore width.

ECOLOGY & DISTRIBUTION — Hypogeous, solitary or in groups in calcareous
soils with pH 6.8-7.6 under Pinus yunnanensis and forming ectomycorrhizae
with its roots, alt. 1500-2000 m, fruiting from autumn to winter. Known only
from Sichuan, China.

ADDITIONAL SPECIMENS EXAMINED: CHINA, SICHUAN PROVINCE, Panzhihua,

26°29’N 102°01’E, under Pinus yunnanensis, 15 Nov. 2012, C.Y. Liu Pan-No-10 (YAAS
L3186; GenBank KJ162155); C.Y. Liu Pan-No-40 (YAAS L3187; GenBank KJ162156).

Discussion

The new Tuber species, T: xanthomonosporum from Sichuan, produces only
one spore per ascus, the diagnostic character of the now defunct genus, Paradoxa
(Mattirolo 1935; Fan et al. 2014). Recent phylogenetic studies of Paradoxa-like
species (Kinoshita et al. 2011; Fan et al. 2014) found that they group with Tuber
species. Our phylogenetic analysis groups T. xanthomonosporum with Tuber
species but not with other Paradoxa-like species, despite sharing the common
character of having only one spore per ascus. Our work therefore supports
the conclusion by Fan et al. (2014) that Paradoxa-like truffle species do not
form a monophyletic group. While Tuber sinomonosporum [= P. sinensis],
T. glabrum, and T. sp. 1 (the Japanese Paradoxa-like new species) do group
together, they also group with T! macrosporum and T. canaliculatum in Clade
I (/macrosporum). These species in Clade I share the same morphological
feature of having red-brown gleba. Tuber xanthomonosporum groups in Clade
II with the /melanosporum and /rufum subclades but with very low bootstrap
support (<50%). Morphologically, T. xanthomonosporum differs greatly from

Tuber xanthomonosporum sp. nov. (China) ... 67

Gymnohydnotrya australiana JQ925629
98 Tuber sp. 1 AB553344
T. sinomonosporum KF002729
100» 7. glabrum KF002731
100 T. glabrum KF002727 Clade I
Tuber sp. 2 AB553356 macrosporum
100) 7. canaliculatum GQ221455
T. canaliculatum GQ221456
100-- 7. macrosporum HM485372
T. macrosporum JF926121
T. xanthomonosporum KJ162154
T. xanthomonosporum KJ162155 Subclade I
T. xanthomonosporum KJ162156
T. indicum JQ638997
T. indicum JQ639005
100 T. indicum JQ639007
T. melanosporum AF167096

100

08 ye T. melanosporum GU810153 Subelade II
T. melanosporum GU810152 melanosporum
100r 7. pseudoexcavatum HM485381 Clade II
100 T. pseudoexcavatum JQ638958

& 100 7. pseudoexcavatum JQ638982

T. pseudoexcavatum JQ639006
T. rufum EF362473
100 T. rufum EF362477
T. rufum JF926123
T. umbilicatum GU979032_ | Subclade TIT
T. umbilicatum GU979031 rufum
go) 2. umbilicatum FJ797879
95! 7. umbilicatum FJ797880
T. borchii AF106890
T. borchii AF250291
83 T. borchii AF132505
100 T. liui DQ898182
T. zhongdianense DQ898186
1001 7, zhongdianense DQ898187
100) 7. latisporum DQ898183
100 T. latisporum DQ898184
109 T. panzhihuanense JQ978644
100! 7. panzhihuanense JQ978650
100) 7. foetidum AJ557543
100 T. foetidum AJ557544
100 7. maculatum AF106889
T. maculatum EU753269
T. excavatum KC330228
loop 7. excavatum HM152020
T. excavatum KC330227
T. fulgens HM152014
100 T. fulgens HM152024
T. fulgens HM485358
100 Choiromyces meandriformis HM485330
Choiromyces meandriformis HM485331

100 100
100

67
100

100

_10

PLATE 2. One of eight most parsimonious trees constructed with ITS sequences of Tuber
xanthomonosporum and related species. MP Bootstrap values greater than 50% are indicated at
nodes.

the species in these two subclades in its whitish to yellow-brownish gleba and
reticulate spores. Given the few species and low support for the species now
placed in Clade I, the phylogenetic relationships need further study with more
samples and gene loci.

Acknowledgements

We are grateful to Dr. Ian Hall and Dr. A. Zambonelli for reviewing the paper. We
also thank Prof. Fan Li and Mr. H.K. Xiong for providing Paradoxa-like specimens for
our study. This study was supported financially by the National Science Foundation
of China (No.31160010) and the Science and technology support program of Sichuan
Province (No.2012NZ0003 and No.2013NZ0029).

68 ... Qing & al.

Literature cited

Chen J, Liu PG. 2007. Tuber latisporum sp. nov. and related taxa, based on morphology and DNA
sequence data. Mycologia 99: 475-481. http://dx.doi.org/10.3852/mycologia.99.3.475

Doyle JJ, Doyle JL. 1987. A rapid DNA isolation procedure from small quantities of fresh leaf
tissues. Phytochemical Bulletin 19: 11-15.

Fan L, Cao JZ, Li Y. 2012. Tuber microsphaerosporum and Paradoxa sinensis spp. nov. Mycotaxon
120: 471-475. http://dx.doi.org/10.5248/120.471

Fan L, Feng S, Cao JZ. 2014. The phylogenetic position of Tuber glabrum sp. nov. and T. sino-
monosporum nom. nov., two Paradoxa-like truffle species from China. Mycological Progress
13(2): 241-246. http://dx.doi.org/10.1007/s11557-013-0908-4

Gardes M, Bruns TD. 1993. ITS primers with enhanced specificity for basidiomycetes-
application to the identification of mycorrhizae and rusts. Molecular Ecology 2(2): 113-118.
http://dx.doi.org/10.1111/j.1365-294X.1993.tb00005.x

Kinoshita A, Sasaki H, Nara K. 2011. Phylogeny and diversity of Japanese truffles (Tuber
spp.) inferred from sequences of four nuclear loci. Mycologia 103(4): 779-794.
http://dx.doi.org/10.3852/10-138

Lzessoe T, Hansen K. 2007. Truffle trouble: what happened to the Tuberales? Mycological Research
111(9): 1075-1099. http://dx.doi.org/10.1016/j.mycres.2007.08.004

Mattirolo O. 1935. Catalogo ragionato dei funghi ipogei raccolti nel Canton Ticino e nelle provincie
Italiane confinanti. Beitrage zur Kryptogamenflora der Schweiz 8: 1-53.

Montecchi A, Sarasini M. 2000. Fungi ipogei d’Europa. A.M.B. Fondazione Centro Studi
Micologici. Vicenza.

Wang Y, Hu HT. 2008. New species of Paradoxa gigantospora comb. nov. from China. Mycotaxon
106: 199-202.

Wang Y, Li ZP. 1991. A new species of Tuber from China. Acta Mycologica Sinica 10(4): 263-265.

White TJ, Bruns T, Lee S, Taylor JW. 1990. Amplification and direct sequencing of fungal ribosomal
RNA genes for phylogenetics. 315-322, in: MA Innis et al. (eds). PCR protocols: a guide to
methods and applications. Academic, New York.

Yang ZL, Zhang LE. 2003. Type studies on Clitocybe macrospora and Xerula furfuracea var. bispora.
Mycotaxon 88: 447-454.

MY COTAXON

http://dx.doi.org/10.5248/130.69
Volume 130, pp. 69-72 January-March 2015

Distance1D - a protein profile analytical program
designed for fungal taxonomy

DuSAN MATERIC??’, BILJANA KUKAVICA}, & JELENA VUKOJEVIC4

'Faculty of Agriculture & Teachers Training Faculty, University of East Sarajevo,
Semberskih ratara bb, 76300 Bijeljina, Bosnia and Herzegovina
*Faculty of Science, The Open University, Walton Hall, MK7 6AA Milton Keynes, UK
*Faculty of Natural Sciences and Mathematics, University of Banja Luka,
Mladena Stojanovica 2, 78000 Banja Luka, Bosnia and Herzegovina
‘Faculty of Biology, University of Belgrade, Takovska 43, 11000 Belgrade, Serbia

* CORRESPONDENCE TO: dusan.materic@gmail.com

ABSTRACT —Taxonomic analysis of macromycete fruiting bodies is a challenging task
that utilizes morphological, biochemical, and molecular methods. Many biochemical and
molecular methods have been developed to test or confirm identifications or phylogenetic
positions independently of morphological data. SDS electrophoresis has been shown to be a
good biochemical method for protein separation. Although protein profiles can be analyzed
by commercially available software, there is no software designed specifically for fungal
taxonomic research. We have developed an open source portable program that uses protein
profiles of fungal fruiting bodies to calculate relative differences between species for use in
generating to generate more accurate phylogenetic trees.

Key worps — biochemistry, fungi, Perl, proteome

Introduction

The challenging task of macromycete taxonomy uses morphological,
biochemical, ecological, and molecular data (Guarro et al. 1999, Blackwell et
al. 2006, Korabecna 2007, Lutzoni & Vilgalys 1995). Sodium dodecyl sulfate
(SDS) electrophoresis is a good biochemical method for protein separation
and biomarker discovery, which when applied to the proteomes of fungal
fruiting bodies gives a good number of separated proteins, revealing small (or
sometimes large) differences in protein expression (Materi¢ 2012). The method,
which is robust and reliable, could serve as an independent taxonomic tool for
fungi (Guarro et al. 1999, Tyrrell 1969).

70 ... Materi¢, Kukavica, & Vukojevi¢é

The aim of this work was to develop a user-friendly open source program
using electrophoresis data for fungal taxonomy. Such a program would compare
and calculate relative differences between taxa based on results from SDS gel
electrophoresis and similar techniques (e.g., western blot, zymography).

Materials & methods

We sampled and identified 21 fruiting bodies representing differently related
species. Proteins were separated according to the method described in Materic (2012),
and protein profiles were obtained for each species. The quality of protein profiles
depends upon many factors during sample homogenization, protein extraction and
electrophoresis, and an optimized protocol can be found in Materic¢ et al. (2012). In this
work we have chosen to show the results of only seven species: however, more detailed
dendrograms are available in Materi¢ (2012).

Protein profile data are stored in .csv format and examples of files can be found along
with the source-code in Materi¢ (2013). Protein profile files include two sets of data:
(1) molecular weights (MW) of proteins; and (2) relative abundance. MW and relative
abundances can be obtained using specialized software such as TotalLab (Phoretix,
Newcastle, United Kingdom). However, abundances should be recalculated as relative
abundances with range of 1 (least abundant) to 5 (most abundant). In order to create
a distance matrix table, protein profiles are compared by distancelD. The distance
matrix tables are transformed into dendrograms using the program package PHYLIP
(Felsenstein 2002).

Results & discussion

Protein electrophoresis, which has been widely used in solving taxonomic and
systematic problems, produces results that are independent of morphological
and molecular data (Guarro et al. 1999, Tyrrell 1969). Thus, it is crucial to
have appropriate software that deals with taxonomic issues rather than with
pure biochemical properties. Apart from distance1D, there is no such software
specifically designed to interpret fungal taxonomy from protein profile data.

Distance1D is a program designed to calculate relative differences between
two protein profiles, which could then be used to generate dendrograms
(Fic. 1). The program is designed and written specifically for use in fungal
taxonomy (mainly for analyzing protein profiles of fungal fruiting bodies), but
its usage could be wider. The program is meant to be open source, user-friendly,
and portable. Portability results from the program's being written in Perl, which
can be run on any operating system. As the program uses the Tk library for
generating widgets (Windows-like interface), Perl/Tk should be installed along
with Perl.

Our previous work produced good protein profiles of fungal fruiting bodies
from SDS gel electrophoresis (Materi¢ 2012) that are stored in separate files
(Fie. 1):

Distance 1D for analyzing fungal profiles ... 71

wW
[ SDS Electrophoresis

Fruiting bodies (Boletus edulis)

distancelD 0.1

Image analysis and storing

Species 1: | /GELS2-3-2Be.lin protein profiles in a .csv files
Species 2: /GELS/2-3-3Sg.lin

TOTAL DIF: 150.7

(mwa):
(abd):

Za > 2 ;
pa Boletus edulis

Boletus luridus

Distance matrix yee
D)

Distances obtained using distance1D

Boletusedulis _|000.0[093.9]217.5] 134.6] 170.0]188.1]150.7

Boletus luridus | 093.9]000.0|182.8]151.0/169.6]1903]161.8) Neighbour joining
Lactarius piperatus — |217.5|182.8/000.0/ 113.0] 133.3]131.0]166.1

Suillus granulatus

Russula cyanoxantha

i
Tree drawing using
i

Lactarius piperatus

Russula foetens

Russula emetica

FIGURE 1. Diagram showing stages of an experiment used in order to gain biochemically based
phylogenetic trees. A: Fruiting bodies. B: Result of SDS electrophoresis (channels: M = molecular
mass marker; 1 = Boletus luridus; 2 = B. edulis; 3 = Suillus granulatus; 4 = Russula foetens;
5 = Lactarius piperatus; 6 = R. cyanoxantha; 7 = R. emetica; Materi¢ et al 2012). C: Interface of
the program distance1D. D: Distance matrix table generated by the program. E: Phylogenetic tree
drawn by PHYLIP program package (similar results were obtained by analyzing ITS II regions of
rDNA as control; Materi¢ 2012).

The program as an input takes the paths of two files (two file names) where
protein data are stored in .csv format. An algorithm searches for optimal pairing
of proteins by looping through the parameters, such as MW sensitivity. After
alignment and optimization, the following differences are scored: (1) presence
of new protein lines (new protein), (2) molecular weight difference of protein
pairs, and (3) abundance difference of protein pairs. The scoring system,
which could be adapted according to user requirements, is set up as follows:
“10” scored for each new protein line, “1” scored for each 1kDa MW shift, and
“2” scored for abundance difference. If DNA/RNA gel data are going to be used,
the abundance difference for each fragment should be set the same.

There are several important advantages to using this program: the program
generates distances that give accurate phylogenetic trees (more accurate than
a single DNA sequence such as rDNA ITS1; Materi¢ 2012); the parameters
can be changed to suit particular research; the program can be easily adapted

72 ... Materié, Kukavica, & Vukojevi¢

for use with any protein/DNA/RNA electrophoresis data; and the program is
suitable for all operating systems and is open source and free.

Acknowledgments
The authors thank Milan Matavulj (University of Novi Sad, Serbia) and Mirjana
Stajic (University of Belgrade, Serbia) for presubmission review.

Literature cited

Blackwell M, Hibbett DS, Taylor JW, Spatafora JW. 2006. Research Coordination Networks: a
phylogeny for kingdom Fungi (Deep Hypha). Mycologia 98: 829-837.
http://dx.doi.org/10.3852/mycologia.98.6.829

Felsenstein, J. 2002. PHYLIP (Phylogeny Inference Package) version 3.6 a3.

Guarro J, Gené J, Stchigel AM. 1999. Developments in fungal taxonomy. Clinical Microbiology
Reviews 12: 454-500.

Korabecna M. 2007. The variability in the fungal ribosomal DNA (ITS1, ITS2, and 5.8S rRNA
gene): its biological meaning and application in medical mycology. 783-787 in Communicating
current research and educational topics and trends in applied microbiology (A Méndez- Vilas,
ed.). Formatex, Spain.

Lutzoni F, Vilgalys R. 1995. Integration of morphological and molecular data sets in estimating
fungal phylogenies. Canadian Journal of Botany 73: 649-659.
http://dx.doi.org/10.1139/b95-307

Materi¢ D. 2012. Biohemijska, molekularna i bioinformaticka analiza taksona gljiva podcarstva
Dikarya. Master's thesis, Bosnia and Herzegovina, University of Banja Luka.

Materi¢ D. 2013. Distance1D - source code. GitHub. https://github.com/dusanac/distance1D

Materi¢ D, Kukavica B, Boroja M, Vukojevic¢ J. 2012. Optimizacija protokola za ekstrakciju proteina
iz plodonosnih tijela gljiva (vrste rodova: Boletus, Russula, Lactarius i Agaricus) za SDS-
elektroforezu. Skup 4(1):36-41.

Tyrrell D. 1969. Biochemical systematics and fungi. Botanical Review 35: 305-316.
http://dx.doi.org/10.1007/BF02858875

MY COTAXON

http://dx.doi.org/10.5248/130.73
Volume 130, pp. 73-77 January-March 2015

Coccomyces prominens sp. nov. (Rhytismataceae)
on Rhododendron coeloneurum in China

YA-FEI Xu’, YUAN Wu’, YAN-QIONG MENG’,
SHI-JUAN WANG?, & YING-REN LIN”

' School of Life Science &? School of Forestry & Landscape Architecture,
Anhui Agricultural University, West Changjiang Road 130, Hefei, Anhui 230036, China

*CORRESPONDENCE TO: yingrenlin@yahoo.com

ABSTRACT — A fungus found on leaves of Rhododendron coeloneurum from Garze prefecture
(Sichuan Province), China, is described as Coccomyces prominens. The new species is similar
to C. urceoloides but differs by its much larger intraepidermal ascomata and the presence of
conidiomata. The type specimen is deposited in the Reference Collection of Forest Fungi of
Anhui Agricultural University, China (AAUF).

Key worps — foliicolous fungus, morphological character, taxonomy, Ericaceae

Introduction

Coccomyces De Not. is the second-largest genus of Rhytismataceae
(Rhytismatales, Leotiomycetes, Ascomycota), represented by 116 species recorded
by Kirk et al. (2008) plus a few recently added. Members of this genus are
widely distributed and occur on leaves, twigs, bark, or wood of vascular plants,
especially Ericaceae, Fagaceae, and Lauraceae (Sherwood 1980). Twenty-eight
Coccomyces species have been reported from China (Korf & Zhuang 1985; Lin
1998; Hou et al. 2006, 2007; Jia et al. 2012; Zheng et al. 2012; Wang et al. 2013;
Yang et al. 2013) since Teng (1934) first recorded C. dentatus (J.C. Schmidt)
Sacc. on Quercus and Castanea and C. delta (Kunze) Sacc. on Lauraceae.

Here, Coccomyces prominens on leaves of Rhododendron coeloneurum from
Luding county, Garze prefecture, Sichuan Province, China, is described as a
new species.

Materials & methods
Mature fruiting bodies were selected from the collected specimens. External shapes,
size, color, ascomatal and conidiomatal openings, and zone lines were observed under

74 ... Xu & al.

a dissecting microscope at 10—-50x magnification. After rehydration in water, 10-20
tum thick vertical transverse sections of fruiting bodies were made using a freezing
microtome and mounted in 0.7% (w/v) cotton blue in water. Gelatinous sheaths
surrounding ascospores and paraphyses were observed in water or 0.1% (w/v) cotton
blue in phenol glycerin. The colors of internal structures and paraphysis contents were
observed in water. Measurements were made using material in 5% KOH and from ca
30 paraphyses, asci, ascospores, conidiogenous cells, and conidia for each specimen.
Figures were drawn of the external and internal structures of the fruiting bodies using a
microscopic drawing tube. The type collection is deposited in the Reference Collection
of Forest Fungi of Anhui Agricultural University, Hefei, China (AAUF).

Taxonomy

Coccomyces prominens Y.F. Xu & Y.R. Lin, sp. nov. Fics 1-6
MycoBank MB 808960
Differs from Coccomyces urceoloides by its much larger, intraepidermal ascomata with

an obvious pre-formed dehiscence mechanism, its wider ascospores enveloped in a
gelatinous sheath, and its production of conidiomata.

Type: China, Sichuan, Garze prefecture, Luding, Hailuogou, alt. ca 3400m, on fallen
leaves of Rhododendron coeloneurum Diels (Ericaceae), 22 June 2009, Y.G. Liu & Y.R. Lin
2768 (Holotype, AAUF 68876).

ETYMOLOGY: prominens (Latin = prominent), referring to the ascomata, which are
noticeable on the substratum.
ZONE LINES usually frequent, brown or dark brown, thin to wide, somewhat
diffused, nearly always partly surrounding the bleached spots.

ConipiomMara in similar positions to ascomata on the substratum, scattered
to clustered, sometimes several coalescent. In surface view, conidiomata
150-220 um diam., rounded or subrounded, with a clearly marked outline,
black-brown, moderately raising the leaf surface, discharging spores through
an apical ostiole. In vertical transverse section, conidiomata intraepidermal,
lenticular to semicircular. UPPER WALL 6.5—12 um thick, almost not attenuating
towards the base, composed of disparately sized angular cells. BASAL WALL
35-50 um thick in the middle, becoming 13-20 um thick at periphery,
consisting of black-brown, angular and elongate, thick-walled cells 3.5-8 um
diam. SUBCONIDIOGENOUS LAYER 5-8 um thick, composed of colorless, thin-
walled angular cells 1.2-2 um diam. CONIDIOGENOUS CELLS 10-14 x 1.5-2.2
um, cylindrical but tapered towards the apex, hyaline, proliferating sympodially
and percurrently. Conrp1a 4.5-7 x 0.5-0.8 um, cylindrical, straight or slightly
curved to one side, ends rounded or bluntly pointed, hyaline, aseptate, smooth-
walled. TRICHOGYNES 35-45 x 2-3 um, cylindrical, tapering towards the apex,
1—2-septate near the base.

Ascomarta epiphyllous, scattered or occasionally coalescent, in subcircular
or irregular pale areas with obvious edges. In surface view, ascomata 800-1400

Fs)

Coccomyces prominens sp. nov. (China) ...

Pee
ory

6. Coccomyces prominens (ex holotype, AAUF 68876) on Rhododendron coeloneurum.

Fics 1

and a zone line observed

ascomata,
under a dissecting microscope. 3. Ascoma in vertical transverse section. 4. Portion of ascoma in

1. A leaf bearing fruiting bodies and zone lines. 2. Conidiomata,

asci, and ascospores. 6. Conidioma in vertical transverse

vertical transverse section. 5. Paraphyses,

section.

76 ... Xu &al.

um diam., quadrilateral to octagonal, black-brown to black, not shiny or
slightly shiny, edge defined, moderately raising the substrate surface, with
an obvious pre-formed dehiscence mechanism, opening by 4-8 radial splits
nearly extend to the edge of ascoma. Lips absent. In median vertical transverse
section, ascomata intraepidermal. COVERING sTROMA 55-65 um thick near
the opening, decreasing to 25-30 um thick adjacent to the base, composed
of black-brown textura angularis-epidermoidea with thick-walled cells 4—6.5
um diam., connecting to the basal stroma. Adjacent to the top of the covering
stroma, a flimsy structure develops consisting of angular, thin-walled, almost
colorless cells staining with cotton blue. BasaL stroma 18-22 um thick,
comprised of 2—3(—4) layers of dark brown, angular, thick-walled cells 4-6 um
diam. ExcrpuLUM well-developed, 45-60 um thick, arising from the marginal
paraphyses and the internal matrix of stroma, consisting of closely agglutinated
septate hyphae 2—5 um diam., mostly short-celled, with swollen greyish-brown
apices. INTERNAL MATRIX OF STROMA moderately developed, 20-50 um thick,
comprising hyaline, gelatinised textura intricata. SUBHYMENIUM colorless,
10-15 um thick, composed of textura angularis. PARAPHYSES 160-190 x
1.8—2.2 um, filiform, 0-1-septate, occasionally branched, yellowish-brown
in the upper quarter, surrounded by a 0.6—0.8 um thick gelatinous matrix,
gradually irregularly swollen to 3-5 um and containing disparately-sized oil
drops near the apex, forming a 35-50 um thick epithecium inserted with a
number of hyaline, refractive, sub-oblong solid gels 12-32 x 8-14 um. AscI
ripening sequentially, 110-170 x 12—15 um, cylindrical-clavate, short-stalked,
thin-walled, apex bluntly pointed to subtruncate-conical, J-, 8-spored.
ASCOSPORES arranged fasciculately, 80-120 x 2-2.4 um, filiform, hyaline,
aseptate, enveloped in a 0.6—0.9 um thick gelatinous sheath.

HOST SPECIES, HABITAT AND DISTRIBUTION: Producing ascomata and
conidiomata on fallen leaves of Rhododendron coeloneurum. Known only from
the type locality, Sichuan Province, China.

COMMENTS — Coccomyces prominens resembles C. urceoloides Spooner in
the well-developed excipulum with more or less brown hyphal tips, gradually
swollen paraphyses at the apex, and refractive solid gels in the epithecium.
However, C. urceoloides has smaller, hypophyllous, subepidermal ascomata
(350-550 um diam.) not associated with conidiomata; much smaller, narrowly
cylindrical asci (120-135 x 5.5—-6 um); and smaller ascospores (80-98 x ca 1
um) with a mucilaginous cap at the proximal end (Spooner 1990).

Coccomyces sinensis Y.R. Lin & Z.Z. Li is easily distinguished from
C. prominens by its triangular to pentagonal ascomata, smaller asci (108-130 x
5.3—-6.0 um), smaller ascospores (54—90 x 1—1.2 um), subhymenium consisting
of textura porrecta-intricata, and distinct paraphyses that are abruptly swollen

Coccomyces prominens sp. nov. (China) ... 77

to subfusoid-ventricose with a subcylindrical mucro near the apex (Lin et al.
2001).

The similar C. monticola Sherwood differs in the smaller and orbicular
ascomata (500-800 um diam.), a thinner covering stroma (20 um thick),
slightly swollen paraphysial apices (2.5 um wide), and much narrower asci (8-9
um wide; Sherwood 1980).

Acknowledgements

We are grateful to Dr J.E. Taylor and Dr M. Ye for serving as pre-submission
reviewers, and to Dr Y.G. Liu for the field investigations. This study was supported by
the National Natural Science Foundation of China (No. 31270065, 31170019).

Literature cited

Hou CL, Piepenbring M. 2007. Two new species of Rhytismataceae on twigs of conifers from
Yunnan Province, China. Mycotaxon 102: 165-170.

Hou CL, Kirschner R, Chen CJ. 2006. A new species and new records of Rhytismatales from
Taiwan. Mycotaxon 95: 71-79.

Jia GJ, Lin YR, Hou CL. 2012 [“2011”]. A new species of Coccomyces (Rhytismatales, Ascomycota)
from Mt. Huangshan, China. Mycotaxon 118: 231-235. http://dx.doi.org/10.5248/118.231

Kirk PM, Cannon PF, Minter DW, Stalpers JA (eds). 2008. Dictionary of the fungi, 10th ed. CAB
International. Wallingford. 771 p.

Korf RP, Zhuang WY. 1985. Some new species and new records of Discomycetes in China.
Mycotaxon 22: 483-514.

Lin YR. 1998. Studies on Coccomyces de Notaris and Neococcomyces gen. nov. in China. [Ph.D.
thesis; in Chinese]. Northeast Forestry University. Harbin. 98 p.

Lin YR, Li ZZ, Xu ZS, Wang JR, Yu SM. 2001. Studies on the genus Coccomyces from China IV
[in Chinese]. Mycosystema 20: 1-7.

Sherwood MA. 1980. Taxonomic studies in the Phacidiales: The genus Coccomyces (Rhytismataceae).
Occasional Papers of the Farlow Herbarium of Cryptogamic Botany 15: 1-120.

Spooner BM. 1990. Coccomyces and Propolis (Rhytismatales) from Mt Kinabalu, Borneo. Kew
Bulletin 45: 451-484. http://dx.doi.org/10.2307/4110513

Teng SC. 1934. Notes on Discomycetes from China. Sinensia 5: 431—465.

Wang SJ, Lin YR, Tang YP, Li K. 2013 [“2012”]. A new species of Coccomyces (Rhytismatales,
Ascomycota) on Ilex elmerrilliana. Mycotaxon 122: 287-291. http://dx.doi.org/10.5248/122.287

Yang MS, Lin YR, Zhang L, Wang XY. 2013 [“2012”]. Coccomyces hubeiensis, a new fungus of
Rhytismatales from China. Mycotaxon 122: 249-253. http://dx.doi.org/10.5248/122.249

Zheng Q, Lin YR, Yu SM, Chen L. 2012 [“2011”]. Species of Rhytismataceae on Lithocarpus spp.
from Mt Huangshan, China. Mycotaxon 118: 311-323. http://dx.doi.org/10.5248/118.311

MY COTAXON

http://dx.doi.org/10.5248/130.79
Volume 130, pp. 79-89 January-March 2015

Type studies on Amauroderma species described by
J.D. Zhao et al. and the phylogeny of species in China

MENG-JIE Li”? & Ha1l-SHENG YUAN”

"State Key Laboratory of Forest and Soil Ecology, Institute of Applied Ecology,
Chinese Academy of Sciences, Shenyang 110164, P. R. China
? University of Chinese Academy of Sciences, Beijing 100049, P. R. China

* CORRESPONDENCE TO: hsyuan@iae.ac.cn

ABSTRACT — Holotypes of eight Amauroderma species described by Ji-Ding Zhao and
co-workers, together with 93 additional specimens collected from China, were examined.
Four names are demoted to synonymy (A. amoiense and A. wuzhishanense = A. rugosum;
A. dayaoshanense = Pyrrhoderma sendaiense; A. fujianense = Ganoderma fornicatum), two
represent accepted species (A. austrosinense, A. yunnanense), and two are of uncertain
application (A. longgangense, A. jiangxiense) because of the poor or sterile condition of the
holotype material. Illustrated descriptions of A. austrosinense and A. yunnanense are given
based on the holotypes. Four other species reported from China (A. perplexum, A. rugosum,
A. subresinosum, A. yunnanense) were sequenced for molecular analysis, and a phylogenetic
tree was constructed by maximum parsimony and Bayesian analyses of the nuclear
internal transcribed spacer of ribosomal DNA (ITS rDNA) sequences. A key to accepted
Amauroderma species in China is provided.

KEY worDs — Ganodermataceae, morphology, phylogenetics, taxonomy

Introduction

Amauroderma Murrill was established by Murrill and typified by A.
regulicolor (Berk. ex Cooke) Murrill (Murrill 1905). This genus is mainly
characterized by sessile or stipitate (mesopodal, pleuropodal) basidiocarps
with a laccate or dull pileus; a trimitic hyphal system; and ellipsoid, subglobose
to globose bitunicate basidiospores with an asperulate inner wall (Furtado
1981, Ryvarden 2004). Macroscopically, Amauroderma shares with Ganoderma
P. Karst. a similar basidiocarp shape of central or lateral stipe and a laccate or
dull surface. Ganoderma can be separated from Amauroderma by its distinctly
truncate basidiospores, and most Ganoderma species grow on dead wood,
while most Amauroderma species grow in the ground from buried roots
(Ryvarden 2004).

80 ... Li & Yuan

Amauroderma is widely distributed in the tropics and subtropics south of
25°N in China within Guangdong, Guangxi, Yunnan, and Hainan Provinces.
Amauroderma species are regarded as economically valuable because of their
famous medicinal properties and pathogenicity (Dai et al. 2007, 2009; Jiao et al.
2013; Chan et al. 2013). Index Fungorum (www.indexfungorum.org) has listed
126 names under this genus. In China, taxonomic research on Amauroderma
began with Teng, who recorded 10 species (Teng 1936, 1939, 1963). Later Zhao
et al. (1979, 1983, 1984) and Zhao & Zhang (1986, 1987a,b, 2000) reported
20 species, nine of which were new. The types of these new species [except
Amauroderma guangxiense J.D. Zhao & X.Q. Zhang (not traced)] are preserved
in the Herbarium Mycologicum Academiae Sinicae (HMAS), the Institute of
Microbiology, Chinese Academy of Sciences. Tai (1979) and Fan & Liu (1990)
have also contributed to Amauroderma records in China.

In this study we investigated the type specimens of Amauroderma described
by Ji-Ding Zhao and his colleagues and 93 other recently collected specimens
from southern China based on morphological characters and rDNA sequence
analyses. The taxa are presented below in alphabetical order according to the
original names.

Materials and Methods

Morphology

The studied specimens were deposited in the Herbaria of the Institute of Applied
Ecology, Chinese Academy of Sciences (IFP); the Institute of Microbiology, Chinese
Academy of Sciences (HMAS); and the Institute of Microbiology, Beijing Forestry
University (BJFC). Microscopic procedures followed He & Dai (2012). Sections were
studied at magnifications up to 1000x using a Nikon Eclipse 80i microscope with phase
contrast illumination. Measurements were made from sections stained with Cotton
Blue. In presenting basidiospore size variation, 5% of measurements are excluded
from each end of the range and given in parentheses. The following abbreviations are
used: CB = Cotton Blue, CB+ = cyanophilous, CB- = acyanophilous, IKI = Melzer’s
reagent, IKI- = negative in Melzer’s reagent, KOH = 5% potassium hydroxide, L = mean
basidiospore length (arithmetic average of all basidiospores), W = mean basidiospore
width (arithmetic average of all basidiospores), Q = variation in the L/W ratios between
the specimens studied, and n = number of basidiospores measured from given number
of specimens. Special color terms follow Petersen (1996).

Molecular phylogeny

Amplification of ITS sequences from herbarium specimens was conducted using
Phire® Plant Direct PCR Kit (Finnzymes, Finland) with ITS5 and ITS4 primers (White et
al. 1990). PCR procedure was as follows: initial denaturation at 98 °C for 5 min, followed
by 39 cycles at 98 °C for 5 s, 59 °C for 10 s and 72 °C for 5 s, and a final extension at
72 °C for 2 min. PCR amplification was confirmed on 1% agarose electrophoresis gels

Amauroderma type studies (China) ... 81

TABLE 1. ITS sequences of Amauroderma and Tomophagus species used in the
molecular analyses.

SPECIES VOUCHER LOCALITY GENBANK No.
A. perplexum Cui 6496 Hainan, China KJ531650
Dai 10811 Hainan, China KJ531651
Wei 5562 Hainan, China KJ531652
A. rugosum Cui 6285 Hainan, China KJ531656
Zhou 153 Guangxi, China KJ531657
Dai 7862 Hainan, China KJ531658
Zhou 136 Guangxi, China KJ531659
Zhou 347 Guangxi, China KJ531660
Zhou 66 Guangxi, China KJ531661
Yuan 4554 Hainan, China KJ531662
Dai 9904 Hainan, China KJ531663
Cui 9011 Guangdong, China KJ531664
Cui 9012 Guangdong, China KJ531665
Wei 5234 Hainan, China KJ531666
Cui 8882 Guangdong, China KJ531667
Dai 9553 Hainan, China KJ531668
Dai 9566 Hainan, China KJ531669
Dai 4345 Hainan, China KJ531670
Dai 10040 Hainan, China KJ531671
Cui 4078 Fujian, China KJ531672
Dai 10307 Hainan, China KJ531673
Zhou 523 Guangxi, China KJ531674
Zhou 547 Guangxi, China KJ531675
Dai 12324 Yunnan, China KJ531676
Dai 12390 Yunnan, China KJ531677
Cui 9006 Guangdong, China KJ531678
THP 30 Unknown HM 480835
A. subresinosum Wei 5569 Hainan, China KJ531649
ML 288 Malaysia JQ409358
A. yunnanense Cui 7974 Yunnan, China KJ531653
Dai 13021 Yunnan, China KJ531654
Yuan 2253 Yunnan, China KJ531655
T. colossus CGMCC 5.763 Philippines JQ081068

stained with ethidium bromide (Stéger et al. 2006). The PCR products were purified and
directly sequenced in Beijing Genomics Institute, China, with the same primers. The
most similar sequences (TABLE 1) were retrieved from GenBank (http://www.ncbi.nlm.
gov) using the BLAST option. Sequences were aligned using ClustalX (Thompson et al.

82 ... Li & Yuan

1997). Alignment was manually adjusted to allow maximum alignment and minimize
gaps, and deposited in TreeBASE (http://treebase.org/treebase-web/; submission ID:
17050). Identity/similarity between two sequences was calculated using the “pairwise
alignment, calculation of the similarity/identity” option of BioEdit v. 7.0.5 (Hall 2005).
Maximum parsimony and Bayesian analysis were applied to the ITS dataset. All
characters were weighted and gaps were treated as missing data. Maximum parsimony
analysis (PAUP* version 4.0b10) was used (Swofford 2002). Trees were inferred using
the heuristic search option with TBR branch swapping and 1,000 random sequence
additions. Max-trees were set to auto-increase, branches of zero length were collapsed,
and all parsimonious trees were saved. Clade stability was assessed using a bootstrap (BT)
analysis with 1,000 replicates (Felsenstein 1985). Descriptive tree statistics, including
tree length (TL), consistency index (CI), retention index (RI), rescaled consistency
index (RC), and homoplasy index (HI) were calculated for all trees generated under
different optimality criteria.

JModelTest (Posada 2008) was used to determine the best-fit evolution model
for each data set for Bayesian inference (BY). Bayesian inference was calculated with
MrBayes3.1.2 with a general time reversible (GTR) model of DNA substitution and a
gamma distribution rate variation across sites (Ronquist & Huelsenbeck 2003). Four
simultaneous Markov chains were run with 3 million generations, and trees were
sampled every 100 generations. Trees generated for the first one-fourth generations
were discarded as burn-in. A 50% majority rule consensus tree of all remaining trees
was calculated. Branches that received maximum parsimony bootstrap support (BSS)
and Bayesian posterior probabilities (BPP) respectively greater than or equal to 75% and
0.95 were considered as significantly supported.

Taxonomy

We studied eight species of Amauroderma (Ganodermataceae) described
from China by Ji-Ding Zhao et al. Four of the eight names are synonymized,
two are accepted, and two are of uncertain application.

Amauroderma amoiense J.D. Zhao & L.W. Hsu, Acta Mycol. Sinica 2: 164. 1983.
Ho.otype: China. Fujian Prov., Xiamen, sandy land, 13.V1.1976, Li Hui-zhong &
Zhang Xiao-qing 28 (HMAS 42784).

= Amauroderma rugosum (Blume & T. Nees) Torrend, Brotéria, Sér. Bot. 18: 127. 1920.
The type material is sterile, but all other characters fit A. rugosum well. For a
detailed description of A. rugosum, see Ryvarden & Johansen (1980).

Amauroderma austrosinense J.D. Zhao & L.W. Hsu, Acta Mycol. Sinica

3: 20. 1984. Fig 1
Hototype: China. Hainan Prov., Bawangling, ground, 20.1V.1977, Han Shu-jin 902
(HMAS 42695).

BASIDIOCARPS annual, stipitate, umbelliform. PILEus single, suborbicular to
circular, glabrous, dull, clay-buff to fulvous brown, densely concentrically zoned

Amauroderma type studies (China) ... 83

Fic. 1. Amauroderma austrosinense (holotype): basidiospores.

and slightly irregularly radially furrowed; MARGIN obtuse and deflexed when
dry; STIPE sub-central or eccentric, concolorous with pileus, cavitate, up to 12
cm long and 1 cm thick. CONTEXT cinnamon, dense and hard, homogenous, up
to 4mm thick. PORE SURFACE cream white when fresh, turning brown with age;
PORES circular, entire, 6-8 per mm. TUBES woody, up to 3 mm long.

HYPHAL SYSTEM trimitic; GENERATIVE HYPHAE with clamp connections,
hyaline, thin-walled, 2.8-4 um in diam; SKELETAL HYPHAE pale yellow to almost
colorless, sparsely branched, 3.7-5.2 um in diam; BINDING HYPHAE hyaline,
branched, tortuous, 1.5-2.2 um in diam; all the hyphae IKI-, CB+; tissues
darkening in KOH. Basipiospores subglobose to globose, faintly yellowish,
IKI-, CB+, doubled-walled, exospore smooth, endospore with conspicuous
echinule, (6.5-)6.8-7.6(-8) x (5.9-)6.2-7(-7.3) um, L = 7.34 um, W = 6.58 um,
Q = 1.12 (n = 30/1).

ADDITIONAL SPECIMEN EXAMINED: CHINA, HAINAN PRov., Changjiang County,

Bawangling Nat. Res., ground, 20.IV.1977 (HMAS 42695).
REMARKS: Amauroderma austrosinense is an accepted species, remarkable for
its umbelliform basidiocarps with distinct concentric rings, small pores, and
basidiospores. A. camerarium (Berk.) J.S. Furtado is similar to A. austrosinense
by sharing a clay-colored, densely concentrically zoned, and radially furrowed
pileus. However, A. camerarium differs in its larger basidiospores (12-15 x
10-13 um; Ryvarden 2004).

Amauroderma leucosporum Corner may be confused with A. austrosinense
by having similar-sized subglobose to globose basidiospores and also not
turning blood red on bruising, but A. leucosporum can be distinguished by its
finely to distinctly villous pileus and stipe (Corner 1983).

84 ... Li & Yuan

Amauroderma dayaoshanense J.D. Zhao & X.Q. Zhang, Acta Mycol. Sinica
6551987
Ho .ortyPe: China. Guangxi Auto. Reg., Dayaoshan, dead wood, 1982, Wei Bing-gang
70 (HMAS 48282).
= Pyrrhoderma sendaiense (Yasuda) Imazeki, Trans. Mycol. Soc. Japan 7: 4. 1966.

The type is sterile but has the characteristics of P sendaiense (see Nunez &
Ryvarden (2000) for a detailed description).

Amauroderma fujianense J.D. Zhao, L.W. Hsu & X.Q. Zhang, Acta Microbiol. Sinica
T9279. 1979:

Ho.otype: China. Fujian Prov., Sanming County, rotten wood, 28.VI.1976, Zhang
Xiao-qing 68 (HMAS 37919).
= Ganoderma fornicatum (Fr.) Pat., Bull. Soc. Mycol. Fr. 5: 71. 1889.
The holotype is sterile but represents a juvenile specimen of G. fornicatum (see
Patouillard (1889) for a detailed description).

Amauroderma jiangxiense J.D. Zhao & X.Q. Zhang, Acta Mycol. Sinica
6: 206. 1987.
Ho .ortyPe: China. Jiangxi Prov., Jian County, rotten wood, 15.11.1983, Yang Ren-gen
152 (HMAS 50418).
The basidiocarp is umbelliform and the pileus is black, laccate, and shiny, but
the specimen is completely sterile. It probably represents a Ganoderma species,
but the application of the name is uncertain.

Amauroderma longgangense J.D. Zhao & X.Q. Zhang, Acta Mycol. Sinica
Sr227 L986,

Ho.otype: China. Guangxi Auto. Reg., Nonggang, on rotten wood, 1979, Wei Bing-
gang 790111-G8 (HMAS 47621).
Basidiospores subglobose, 6-8.5 x 5-8 um, with spinules 0.8-1.2 um on
sporoderm. However, the single-walled basidiospore obviously differs from
the double-walled spore of other Amauroderma species. The application of the
name is uncertain.

Amauroderma wuzhishanense J.D. Zhao & X.Q. Zhang, Acta Mycol. Sinica
6: 208. 1987.

Ho.otype: China. Hainan Prov., Wuzhishan, on rotten wood, 22.VII.1956, Jiang
Guang-zheng (HMAS 19311).

= Amauroderma rugosum (Blume & T. Nees) Torrend, Brotéria, Sér. Bot. 18: 127. 1920.

The holotype is in accordance with Amauroderma rugosum, with its pileus
obscurely zonate, radially quite rugose, and warted. For a detailed description
of A. rugosum, see Ryvarden & Johansen (1980).

Amauroderma type studies (China) ... 85

Fic. 2. Amauroderma yunnanense (holotype): basidiospores.

Amauroderma yunnanense J.D. Zhao & X.Q. Zhang, Acta Mycol. Sinica,
Suppl. 1: 268. 1987 [“1986”]. FIG 2

Ho.otype: China. Yunnan Prov., Xichou County, ground, Xiaogiaogou, 14.V.1959,

Wang Qing-zhi 89 (HMAS 48231).
BASIDIOCARPS annual, centrally to laterally stipitate, coriaceous to corky,
brittle when dry. PiLeus single, flabelliform, suborbicular, umbilicate, PILEAL
SURFACE pale yellowish brown to dark brown, adpressed velutinate, with distinct
concentric zones at the margin and fine furrows in the middle; MARGIN inflexed,
acute or subacute, entire or slightly lacerated and thin; stipe concolorous with
the pileus, up to 7 cm long, inflated at base. ConTExT homogeneous, white
to pale yellow. PoRE SURFACE white when fresh, pale straw yellow when dry,
bruising unchanged; porgs angular, 2-3 per mm; TuBEs concolorous with pore
surface, up to 5 mm thick.

HYPHAL SYSTEM trimitic; GENERATIVE HYPHAE with clamps, hyaline,
thin-walled, 2-4.3 um in diam; SKELETAL HYPHAE thick-walled, pale yellow,
arboriform, 3.5-5.5 um in diam; BINDING HYPHAE hyaline, branched, tortuous,
1.8-2.5 um in diam; all the hyphae IKI-, CB+; tissues darkening in KOH.
Basipiospores broadly ellipsoid to subglobose, pale yellow brown, IKI-,
CB+, doubled-walled, exospore smooth, hyaline, endospore with conspicuous
echinule, (8.3—)8.9-10.3(-10.7) x (6.3-)6.7-8(-8.8) um, L = 9.35 um, W = 7.32
um, Q = 1.22-1.34 (n = 120/4).

ADDITIONAL SPECIMENS EXAMINED: CHINA, YUNNAN PRov., Chuxiong, Zixishan
Forest Park, ground, 8.[X.2006, Yuan 2253 (IFP 013207); Ailaoshan, 5.[X.2007, Yuan
3367 (IFP 013182); Kunming, Qiongzhusi Park, 21.X.2009, Cui 7974 (BJFC 006463);
Xiaoshao Village, 22.1X.2012, Dai 13021 (IFP 019123).
REMARKS: Amauroderma yunnanense is an accepted species characterized by a
brown tomentose pileus and stipe.

Amauroderma perplexum Corner and A. yunnanense both have a velutinate

pileus and stipe as well as stipes that dilate at the base; A. perplexum can be

86 ... Li & Yuan

separated by larger basidiospores, longer tubes, and a pore surface that changes
color when bruised (Corner 1983).

Amauroderma partitum (Berk.) Wakef., which may be confused with
A. yunnanense as they share similar angular pores that are 2-3 per mm, can
be distinguished by its glabrous, slightly shiny pileus and larger basidiospores
(12-15 x 8-10 um; Ryvarden 2004).

Molecular phylogeny

The nuclear ITS rDNA dataset contains 32 sequences representing
A. perplexum, A. rugosum, A. subresinosum (Murrill) Corner, and A. yunnanense,
with Tomophagus colossus (Fr.) Murrill as outgroup (TasBLe 1). After the
ambiguous sites at both ends and the 5.8S region were trimmed off, the
alignment comprised 561 characters, of which 462 were constant, 22 were
variable but parsimony-uninformative, and 77 were parsimony-informative.
Maximum parsimony analysis yielded 6 equally parsimonious trees
(CI = 0.918, RI = 0.952, RC = 0.874, HI = 0.082). A strict consensus tree of these
trees is presented in Figure 3. The 50% majority consensus tree generated by
the Bayesian analysis showed a similar topology with the strict consensus MP
tree, and only the topology from MP analysis is presented, while both bootstrap
support value (BSS) and Bayesian posterior probabilities (BPP) are shown at
the nodes (Fic. 3).

In the phylogenetic tree, each sampled Amauroderma species was resolved
as a strongly supported clade (100% BSS and 1.00 BPP). Identity/similarity
calculation result shows that identities of Amauroderma sequences are
between 0.98 and 1. We failed to obtain sequences from the type specimens of
Amauroderma described by Ji- Ding Zhao from China as the materials were too
old to extract integrated DNA.

Key to species of Amauroderma in China

i. Pileal surface appressed welutinate:.o iiss talee ge hues Aol h eva heh Wale Thee 2
Te Pileal ‘surface: PlaDEOUS.§ 4p seven aybscton teh acbea dring.seg rhea son dhe soa dle puon toby baud he fobs deh fro 9 3
2. Pores 2-3 per mm, pore surface straw yellow, not reddening on bruising,

spores (8.3—)8.9-10.3(-10.7) x (6.3-)6.7-8(-8.8) um. .......... A. yunnanense
2. Pores 4-6 per mm, pore surface cream white, reddening on bruising,

spores (10.3-)10.8-14(-14.8) x (8.9-)9.2-12(-13.1) um......... A. perplexum
3. Pileus surface laccate, spores ovoid to broadly ellipsoid,

(13.7-)14.6-17.1(-17.5) x (8.3-)8.9-10.2(-11.3) um.......... A. subresinosum
SaPileal Sut acevcUlle., £ reste dove. tdan ove len aortas Sahy eeselger estes ace aeearatenat hts gt mmaie ates -:
4, Pileal clay-buff to fulvous brown, pores 7-8 per mm,

spores (6.5—)6.8-7.6(-8) x (5.9-)6.2-7(-7.3) um .........0.00. A. austrosinense

4. Pileus taupe, dark brown to black, pores 4-6 per mm,
spores (8.1—)8.4-11.7 (-12) x (7-)7.5-9.6(-10.3) um. ............. A. rugosum

Amauroderma type studies (China) ... 87

100/1.00 rAmauroderma subresinosum Wei 5569
Amauroderma subresinosum JQ 409358
Amauroderma yunnanense Cui 7974
ROUEOe Amauroderma yunnanense Dai 13021
100/1.00 Amauroderma yunnanense Yuan 2253
Amauroderma perplexum Cui 6496
100/08 Amauroderma perplexum Wei 5562
‘Amauroderma perplexum Dai 10811
Amauroderma rugosum Cui 6285
52/0.52 Amauroderma rugosum Zhou 153
Amauroderma rugosum Dai 7862
Amauroderma rugosum Zhou 136
Amauroderma rugosum Zhou 347
Amauroderma rugosum Zhou 66
62/0.97 Amauroderma rugosum Yuan 4554
Amauroderma rugosum Dai 9904
Amauroderma rugosum HM 480835
Amauroderma rugosum Cui 9011
Amauroderma rugosum Cui 9012
Amauroderma rugosum Wei 5234
Amauroderma rugosum Cui 8882
100/1.00 Amauroderma rugosum Dat 9553
Amauroderma rugosum Dai 9566
Amauroderma rugosum Dai 4345
Amauroderma rugosum Dai 10040
Amauroderma rugosum Cui 4078
Amauroderma rugosum Dai 10307
Amauroderma rugosum Zhou 523
Amauroderma rugosum Zhou 547
Amauroderma rugosum Dai 12324
Amauroderma rugosum Dai 12390
Amauroderma rugosum Cui 9006
Tomophagus colossus JQ 081068
Liooorsiets
0.0 8.0

Fic. 3. Strict consensus tree obtained from Maximum Parsimony and Bayesian analyses of ITS
sequences of Amauroderma. Tomophagus colossus was used as outgroup. Numbers at branches
indicate parsimony bootstrap values and Bayesian posterior probabilities values higher than 50%.

88 ... Li & Yuan

Acknowledgements

The authors are grateful for presubmission review and helpful comments by Bao-
Kai Cui (Institute of Microbiology, Beijing Forestry University) and Xiao-Yong Liu
(Institute of Microbiology, Chinese Academy of Sciences). Special thanks are due to Dr.
Katherine F. LoBuglio (Harvard University Herbaria, USA) for checking and improving
the manuscript. This research was financed by the National Natural Science Foundation
of China (Project Nos. 31170022 & 31470148).

Literature cited

Chan PM, Kanagasabapathy G, Tan YS, Sabaratnam V, Kuppusamy UR. 2013. Amauroderma
rugosum (Blume & T. Nees) Torrend: nutritional composition and antioxidant and potential
anti-inflammatory properties. Evid.-Based Compl. Alt. Med. 2013, Article ID 304713, 10 pages.
http://dx.doi.org/10.1155/2013/304713

Corner EJH. 1983. Ad Polyporaceas I. Amauroderma and Ganoderma. Beheift. Nova Hedwigia 75.
183 p.

Dai YC, Cui BK, Yuan HS, Li BD. 2007. Pathogenic wood-decaying fungi in China. Forest Pathol.
37:105-120. http://dx.doi.org/10.1111/j.1439-0329.2007.00485.x

Dai YC, Yang ZL, Cui BK, Yu CJ, Zhou LW. 2009. Species diversity and utilization of medicinal
mushrooms and fungi in China. Int. J. Med. Mushr. 11: 287-302.
http://dx.doi.org/10.1615/Int)MedMushr.v11.i3.80

Fan L, Liu B. 1990. A new species of Amauroderma. Acta Micol. Sinica 9: 202-205.

Felsenstein J. 1985. Confidence intervals on phylogenetics: an approach using bootstrap. Evolution
39: 783-791.

Furtado JS. 1981. Taxonomy of Amauroderma (Basidiomycetes, Polyporaceae). Mem. New York
Bot. Gard. 34. 109 p.

Hall T. 2005. BioEdit: biological sequence alignment editor for Win95/98/NT/2 K/XP. Ibis
Therapeutic, Carlsbad, CA.

He SH, Dai YC. 2012. Taxonomy and phylogeny of Hymenochaete and allied genera of
Hymenochaetaceae (Basidiomycota) in China. Fungal Divers. 56: 77-93.

Jiao CW, Xie YZ, Yang XL, Li HR, Li XM, Pan HH, Cai MH, Zhong HM, Yang BB. 2013. Anticancer
activity of Amauroderma rude. PLOS ONE 8(6): e66504.
http://dx.doi.org/10.1371/journal.pone.0066504

Murrill MA. 1905. The Polyporaceae of North America XI. A synopsis of the brown pileate species.
Bull. Torrey Bot. Club 32: 353-371.

Nujez M, Ryvarden L. 2000. East Asian polypores. Synopsis Fungorum 13. 168 p.

Patouillard NT. 1889. Le genre Ganoderma. Bulletin de la Société Mycologique de France 5: 64-80.

Petersen JH. 1996. Farvekort. The Danish Mycological Society’s colour-chart. Foreningen til
Svampekundskabens Fremme, Greve.

Posada D. 2008. jModelTest: Phylogenetic model averaging. Mol. Biol. Evol. 25: 1253-1256.
http://dx.doi.org/10.1093/molbev/msn083

Ronquist F, Huelsenbeck JP. 2003. MRBAYES 3: Bayesian phylogenetic inference under mixed
models. Bioinformatics 19: 1572-1574. http://dx.doi.org/10.1093/bioinformatics/btg180

Ryvarden L. 2004. Neotropical polypores Part 1. Syn. Fung. 19: 38-69.

Ryvarden L, Johansen I. 1980. A preliminary polypore flora of East Africa. Fungiflora, Oslo.

Stdger A, Schaffer J, RuppitschW. 2006. A rapid and sensitive method for direct detection of Erwinia
amylovora in symptomatic and asymptomatic plant tissues by polymerase chain reaction.
J. Phytopathol. 154: 469-473.

Amauroderma type studies (China) ... 89

Swofford DL. 2002. PAUP*. Phylogenetic analysis using parsimony (*and other methods). Version
4.0b10. Sinauer Associates, Sunderland, Massachusetts.

Tai FL. 1979. Sylloge Fungorum Sinicorum. Science Press, Peking (in Chinese).

Teng SC. 1936. Additional fungi from China III. Sinensia. 7:529-569.

Teng SC. 1939. A contribution to our knowledge of the higher fungi of China. National Institute of
Zoology & Botany, Academia Sinica.

Teng SC. 1963. Fungi of China. Science Press, Beijing (in Chinese).

Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG. 1997. The Clustal_X windows
interface: flexible strategies for multiple sequence alignment aided by quality analysis tools.
Nucleic Acids Res. 25:876-4882.

White TJ, Bruns T, Lee S, Taylor J. 1990. Amplification and direct sequencing of fungal ribosomal
RNA genes for phylogenetics. 315-322, in: MA Innis MA et al. (eds). PCR protocols, a guide to
methods and applications. Academic, San Diego.

Zhao JD, Zhang XQ. 1986. Taxonomic studies on Ganodermataceae of China V. Acta Mycol. Sinica
Br 219F2 25;

Zhao JD, Zhang XQ. 1987a [“1986”]. Taxonomic studies on Ganodermataceae of China VII - the
genus Amauroderma. Acta Mycol. Sinica, Suppl. 1: 258-272.

Zhao JD, Zhang XQ. 1987b. Taxonomic studies on Ganodermataceae of China VII. Acta Mycol.
Sinica 6: 199-210.

Zhao JD, Zhang XQ. 2000. Flora Fungorum Sinicorum, vol. 18. Ganodermataceae (in Chinese).
Sciences Press, Beijing.

Zhao JD, Xu LW, Zhang XQ. 1979. Taxonomic studies on the subfamily Ganodermoideae of China
(in Chinese). Acta Microbiol. Sinica 19: 265-279.

Zhao JD, Xu LW, Zhang XQ. 1983. Taxonomic studies on Ganodermataceae of China II. Acta
Mycol. Sinica 2: 159-167.

Zhao JD, Xu LW, Zhang XQ. 1984. Taxonomic studies on Ganodermataceae of China III. Acta
Mycol. Sinica 3: 15-23.

MY COTAXON

http://dx.doi.org/10.5248/130.91
Volume 130, pp. 91-103 January-March 2015

Cladonia corymbescens consists of two species

TEUVO AHTI, RAQUEL PINO-BODAS, & SOILI STENROOS

Botanical Museum, Finnish Museum of Natural History, University of Helsinki,
PO. Box 7, FI-00014 Helsinki, Finland

* CORRESPONDENCE TO: teuvo.ahti@helsinki.fi

Asstract — The lichen Cladonia corymbescens (Cladoniaceae, Ascomycota) has been
reported to have two main ranges, one in Melanesia and Australasia and another in the
Himalayas and surrounding areas. When specimens from New Caledonia, Bhutan, and
Thailand were subjected to molecular sequencing (ITS rDNA and RPB2), two distinct clades
were detected. The clades were also supported by morphological and geographical differences
supporting recognition of two distinct species. Cladonia corymbescens s. str. is present in
the Philippines, Melanesia, and Australasia, while the South East Asian mainland material is
referred to C. pseudofissa, a new combination at species level.

Key worps — Cladonia furcata group, lichens, molecular systematic, phylogeny, taxonomy

Introduction

One of the most common fruticose species of Cladonia in the mountains of
western Melanesia is Cladonia corymbescens. The nomenclature of this species
and its similarities with other species was thoroughly discussed by Stenroos
(1988), who also reported it from China and Nepal. However, Abbayes (1974)
was the first lichenologist to recognize this species from the Himalayas. His
opinion was also accepted by Awasthi & Ahti (2007) and Rai et al. (2014), who
stated that the species is widespread in the mountains in India and Nepal. In
his world monograph, Vainio (1887) could not give a definite opinion on the
type material of C. corymbescens from New Caledonia, and he referred the only
Himalayan specimen representing that species that he apparently examined to
C. erythrosperma var. thomsonii.

Several studies using molecular data have shown that many morphological
Cladonia species are not monophyletic (Kotelko & Piercey-Normore 2010,
Fontaine et al. 2011, Pino-Bodas et al. 2011, Steinova et al. 2013). The re-
examination of the morphology based on molecular data in lichenized fungi
has revealed that in several cases some phenotypical differences have been
overlooked (Argitiello et al. 2007, Nufez-Zapata et al. 2010, Parnmen et al.

92 ... Ahti, Pino-Bodas, & Stenroos

2013, Muggia et al. 2014). The aim of this study was to use DNA sequence
data to assess whether Cladonia corymbescens is a genetically homogeneous
species and to revise its classification based on the phylogenetic results and
morphological differences.

Materials & methods

Material for DNA analyses

Altogether 39 specimens of 29 species of Cladonia representing the supergroup
Cladonia in the provisional classification of Stenroos et al. (2002) were selected for a
phylogenetic analysis in this study. These specimens are preserved in the herbaria of
University of Helsinki (H), University of Turku (TUR) or Complutense University,
Biology Faculty (MACB); details of the collections are shown in TaBLE 1. Four specimens
of Cladonia corymbescens, two from New Caledonia and two from the Himalayas, were
available for DNA analyses, while 60 sequences had been generated in previous studies
(Stenroos et al. 2002; Pino-Bodas et al. 2010a, 2010b, 2012a, 2012b, 2012c, 2013a,
2013b). Cladonia rangiformis Hoffm. was selected as outgroup based on previous results
(Stenroos et al. 2002).

DNA extraction, PCR, and sequencing

Total DNA was extracted using DNeasy Plant Mini Kit (QUIAGEN, Germany)
following the manufacturer's instructions. In this study two genetic regions were
chosen: internal transcribed spacer of ribosomal DNA (ITS rDNA) and the second
largest subunit of RNA polymerase IT (RPB2). Although the ITS rDNA region has
been chosen as standard barcoding for fungi (Schoch et al. 2012), in Cladonia the
ITS rDNA frequently fails in the identifications (Kelly et al. 2011; Pino-Bodas et al.
2013b) and RPB2 gives better results (Pino-Bodas et al. 2013b). The primers used to
amplify the nuclear ITS nrDNA were ITS1F (Gardes & Bruns 1993) and ITS4 (White
et al. 1990), and for RPB2 they were RPB2dRaq/RPB2rRaq (Pino-Bodas et al. 2010a)
or CLRPB25F/CLRPB27R (Yahr et al. 2006). The amplification programs are listed in
Pino-Bodas et al. (2013a). PCR was carried out with Ready-to-Go-PCR Beads (GE
Healthcare Life Sciences, UK). Amplifications were prepared for a 25 uL final volume.
PCR was performed using a Mastercycler ep Gradient S (Eppendorf, Westbury, NY).
PCR products were purified with ExoSAP-IT (USB Corporation, USA) and sequenced
in Macrogen Europe (Amsterdam, Netherlands).

Alignments and phylogenetic analyses

The forward and reverse strands were assembled and edited in Sequencher™ v. 4.1.4
(Gene Codes Corporation, Inc., Ann Arbor, Michigan, USA). BLAST searches were
used to confirm that the obtained sequences belonged to Cladonia. The sequences were
manually aligned in BIOEDIT (Hall 1999). Nine ambiguous positions were manually
delimited and removed from the ITS rDNA alignment. The RPB2 alignment did not
have ambiguous positions.

Every region was separately analyzed by Maximum Likelihood (ML), with 500
bootstrap replicates to assess the support of each node. The conflict between the loci
was tested according Lutzoni et al. (2004). This method considers the existence of

Cladonia pseudofissa comb. nov. (Southeast Asia) ... 93

TABLE 1. List of Cladonia specimens used in the molecular study with voucher
specimen information and GenBank accession numbers.
(The new sequences and the new combination are in bold font.)

SPECIES COLLECTION CODE ITS rDNA RPB2
C. acuminata USA, Ahti 63278 (H) 1ACUMI JN621932 JN621965
C. apodocarpa USA, Harris 54250 (H) 1APODO KC526127 KC526068
C. borbonica New Caledonia, Dennetiére 109 (H) AT686 AF455214 KP732364
C. cariosa Spain, Burgaz (MACB 94208) 5CARI JN621909 JN621941
C. cartilaginea Brazil, Stenroos 4926 (TUR) LK45 AF455212 —
C. ceratophylla Brazil, Stenroos 5081 (TUR) LK37 AF455171 —
C. conista Spain, Burgaz (MACB 92796) 2HUMIL JF926613 JF926567
Russia, Zolotuchin 26a (H) 1CONIST JF926633 JF926568
C. corsicana Spain, Burgaz (MACB 100763) SP1 JE288797 JF288833
Spain, Burgaz (MACB 101074) SP2 JF288798 JF288834
C. corymbescens New Caledonia, Dennetiére 45 (TUR) AT680 AF455235 KP732366
New Caledonia, Christenhusz 6169d (H) CL166 KP732363 KP732367
C. cyathomorpha Spain, Burgaz (MACB) 1CYATH KC415941 KC525275
Norway, Ahti 68660 & Tonsberg (H) 3CYATH KC415943 = KC525276
C. firma Spain, Burgaz (MACB 91619) 1FIRM FM205907 FM207568
Spain, Burgaz (MACB 90655) 7FIRM FM205910 FM207576
C. foliacea Portugal, Burgaz (MACB 90506) 1FOL FM205894 FM207569
C. furcata USA, Ahti 58283 (TUR) AT638 AF455220 KP732369
C. humilis Spain, Burgaz (MACB 95931) 9HUMIL JF926615 JF926576
Croatia, Burgaz (MACB 101103) 20HUMIL JF926621 JF926580
C. multiformis Canada, Ahti 57065 (H) LK70 AF455213 KP732370
C. nana Brazil, Stenroos 4940 (TUR) LK34 AF455211 —
C. neozelandica New Zealand, Wirth 28180 (TUR) AT590 AF455206 —
C. petrophila USA, Ahti 56654 (H) LK68 AF455222. —
C. peziziformis USA, Stenroos 5198 (TUR) AT631 AF455221 —
USA, Ahti 56670 (H) LK38 AF455182 —_
C. pityrophylla Brazil, Stenroos 5096 (TUR) LK40 AF455238 —
C. pseudofissa Thailand, Parnmen 285 (H) 1CORYMB KP732362 KP732365
Bhutan, Sgchting 9206 (H) LK35 AF455239 KP732368
C. pulvinata Spain, Burgaz (MACB 91646) 4PUL FM205911 =FM207579
C. pulvinella USA, Ahti 69191 (H) CLCAL4 KC415963 KC525287
C. pyxidata Greenland, Hansen (H) 17PYXI KC415983 = KC525269
C. rangiformis Netherlands, Van der Goes et al. (H) 5RANG JN811400 JN811429
Sweden, Skytén (H) 6RANG JN811401 JN811430
C. scabriuscula Canada, Ahti 56969 (H) LK11 AF455217 KP732371
C. subcariosa USA, Moore (H) 1SUBCARI JN621936 JN621969
C. subconistea Korea, Moon 7188 (H) 1SUBCONI KC415949 KC525296
C. subulata Spain, Burgaz (MACB 93151) 1SUBU FN86566 HM243210
C. turgida Canada, Lendemer (H) 1TURG JF288801 KC526089

94 ... Ahti, Pino-Bodas, & Stenroos

incongruity whenever a clade was supported with a bootstrap of more than 75% in a
locus, while in other loci the individual sequences of this clade were part of another
clade with bootstrap support =75%. The trees were checked manually. No conflicts
were detected and the datasets were combined. The combined dataset was analyzed by
Maximum Parsimony (MP), ML, and Bayesian analyses. MP analysis employed PAUP
version 4.0.b.10 (Swofford 2002), using heuristic searches with 1000 random taxon-
addition replicates with TBR branch swapping and MulTrees option in effect, and equally
weighted characters and gaps treated as missing data. Bootstrap with 1000 replicates
was used as the confidence analysis, using the heuristic option. The ML and Bayesian
analyses were run considering four partitions (ITS rDNA and each codon position of
RPB2). Maximum Likelihood analyses (ML) were done in RAXML 7.04 (Stamatakis
2006), assuming the model GTRGAMMA. A fast bootstrap with 500 replicates was
implemented to assess the support of each node. Bayesian analysis was carried out using
the program MrBayes 3.1.2 (Huelsenbeck & Ronquist 2001). The models of evolution
were selected with MrModeltest (Nylander 2004) under AIC criterion. The models
selected were GTR+I+G for ITS rDNA and SYM+1+G for RPB2. Posterior probabilities
(pp), approximated by sampling trees using Markov Chain Monte Carlo (MCMC), of
each branch were calculated by counting the frequency of trees that were visited during
the course of the MCMC analysis. Model parameters were estimated in each analysis for
5,000,000 generations sampled in four simultaneous chains, and every 1000th was saved
into a file. When the analysis was finished we checked that standard deviation between
the runs was <0.005. The program AWTY (Nylander et al. 2008) was used to determine
when the chains reached the stationary stage. The first 1000 trees were deleted as the
“burn in” and the 50% majority-rule consensus tree was calculated using the “sumt”
command of MrBayes.

Morphology and chemistry

About 63 herbarium specimens of Cladonia corymbescens s. lat. deposited in Helsinki
(H) were morphologically re-evaluated after the phylogenetic analyses. Authors Ahti
and Stenroos had previously studied many additional specimens in various herbaria
[e.g., Geneva (G), Leiden (L), Lucknow (LWG, LWG-LWU, LWG-AWAS)] as well as
material borrowed from Edinburgh (E); see also the specimen lists in Stenroos (1988)
and Rai et al. (2014).

The secondary metabolites were analyzed by thin layer chromatography (TLC)
according to standardized procedures (Orange et al. 2001) using the solvents A (Toluene:
dioxane: acetic acid, MERCK) and B (Hexane: diethylether: formic acid, MERCK).

Results

In this study we generated 10 new sequences (2 of ITS rDNA and 8 of RPB2).
The combined dataset contained 1332 unambiguous characters (597 of ITS
rDNA and 735 of RPB2), 267 of which were parsimony informative (152 of ITS
rDNA and 115 of RPB2). The MP analysis generated two equally parsimonious
trees of 886 steps. The ML analyses of the combined dataset generated a tree
with likelihood value of -LnL = 6355.82, while the mean of likelihood of the
Bayesian analysis was —LnL = 6589.51.

Cladonia pseudofissa comb. nov. (Southeast Asia) ... 95

77 --C. apodocarpa 1APODO
C. petrophila LK68
C. borbonica AT686
100"— ©. corsicana SP 1
C. corsicana SP2
1por— C. cyathomorpha 1CYATH
C. cyathomorpha 3CYATH
C. conista 1CONIST
C. conista 2HUMIL
C. subconistea 1SUBCONI
C. pulvinella CLCAL4
9gr— C. cartilaginea LK45
87 C. neozelandica AT590
85 C. nana LK34
100(— C. peziziformis AT631
C. peziziformis LK38
C. furcata AT638
C. multiformis LK70
100 C. humilis 2OHUMIL
C. humilis 9HUMIL
C. scabriuscula LK11
100[— ©. corymbescens AT680 (New Caledonia)
C. corymbescens CL166 (New Caledonia)
gi1-— C. ceratophylla LK37 A
C. pityrophylla LK40
1por— C. pseudofissa comb. nov. 1CORYMB (Thailand)
C. pseudofissa comb. nov. LK35 (Bhutan)
100/— ©. firma 1FIRM
C. firma 7FIRM
78[—- C. acuminata 1ACUMI
C. cariosa 5CARI
C. foliacea 1FOL

100

100 g9f— C. pulvinata 4PUL
95 C. subcariosa 1SUBCARI
le C. subulata 1SUBU

C. turgida 1TURG
C. pyxidata 17PYXI|
C. rangiformis 5RANG
C. rangiformis BRANG

Fic. 1. Cladonia spp. phylogeny: consensus tree of the two most parsimonious trees based on the
concatenated matrix of ITS rDNA and RPB2. The numbers on the branches represent bootstrap
values (>75%).

In all the analyses the specimens from the Himalayan region and the
specimens from New Caledonia formed two clades; however, the relationships

96 ... Ahti, Pino-Bodas, & Stenroos

om Lome C. porbonics RvO8S
¢. corsicana
74 re ose i fe CYATH
0.99/82] 1/100 . cyatnomorpna
C. cyathomorpha 3CYATH_

1/100 C. cartilaginea LK45
-_C, neozelandica AT590
C. nana LK34 .

C. peziziformis AT631
C. peziziformis LK38
C. furcata AT638
C. multiformis LK70
1/100 - C. humilis ZOHUMIL
C. humilis 9HUMIL
0.97/77 1/100 -C. conista 1CONIST
1/88 C. conista 2HUMIL
C. subconistea_1SUBCONI

C. pulvinella CLCAL4

C. scabriuscula

1
1/88 C. ceratophylla LK37
ane eee aes eS pityrophylla LK40

C. pseudofissa comb. nov. 1CORYMB (Thailand)
C. pseudofissa comb. nov. LK35 en
1/100; C. corymbescens AT680 (New Caledonia

C. corymbescens CL166 (New Caledonia)
al Se eC apodocarpa 1APODO
C. petrophila LK6

C. pulvinata 4PUL
C. subcariosa 1SUBCARI

C. subulata 1SUBU
- C. turgida 1TURG
C. acuminata 1ACUMI
C. cariosa 5CARI
14100 - C. rangiformis SRANG

C. rangiformis BRANG

Fic. 2. Cladonia spp. phylogeny: 50% consensus majority tree of Bayesian analysis based on the
concatenated matrix of ITS rDNA and RPB2. Posterior probability of bayesian analysis (>0.95) and
bootstrap values of ML analyses (>75%) are shown on the branches.

were not the same in all the analyses. In the MP analysis (Fie. 1) the specimens
from the Himalayan region (in the tree C. pseudofissa) are closely related with
C. pityrophylla Nyl. and C. ceratophylla (Sw.) Spreng. (Fic. 1, clade A), but this
relationship was not well supported (bootstrap <70%). The specimens from
New Caledonia cluster in a clade with specimens representing the C. furcata
group, C. humilis group, C. corsicana (Rondon & Vézda) Pino-Bodas et al.,
C. borbonica Nyl., C. cartilaginea Mill. Arg., and C. neozelandica Vain. (Fie. 1,
clade B). The ML and Bayesian analyses generated trees with the same topology,
and only the 50% consensus majority-rule tree from Bayesian analyses is
shown (Fic. 2). The specimens from New Caledonia (C. corymbescens) and the
specimens from the Himalayan region (C. pseudofissa) formed two clades that
are closely related, although the relationship was not supported in any of the
analyses.

Other differences between the MP tree and the Bayesian tree are that
C. pyxidata (L.) Hoffm. was shown as basal in the MP analysis, while the
Bayesian analysis showed C. cariosa (Ach.) Spreng. and C. acuminata (Ach.)
Norrl. in the basal clade. Additionally, the clade of C. conista (Nyl.) Robbins,

Cladonia pseudofissa comb. nov. (Southeast Asia) ... 97

C. subconistea Asahina, and C. pulvinella S. Hammer is closely related to a clade
of C. cyathomorpha Stirt. ex Walt. Watson, C. corsicana, and C. borbonica in the
MP analysis, but not in the Bayesian analysis.

Taxonomy

Cladonia corymbescens Ny]. ex Leight., Ann. Mag. Nat. Hist., ser. 3, 18: 407.
1866. Fig. 3A

TyPeE (Abbayes 1974: 112): New Caledonia, Mont de M’bée, 1855-1860 E. Vieillard 1785
(PC, lectotype, G, H-NYL 38414, PC-Lenormand, isolectotypes). [The lectotypification
by Abbayes was overlooked by Ahti in Stenroos (1988: 126), Ahti (1993) and other
recent authors, but the type collection cited is the same].

= Cladonia novoguineensis Zahlbr., Bot. Jahrb. Syst. 62: 455. 1929.

Type: Papua New Guinea [“Nordéstliches Neu-Guinea, Kaiser Wilhelmsland”], Morobe

Prov., Sarawaged Mts. [“Saruwaged-Gebirg”], 3600-4000 m, C. Keysser 66 [“68”] (W,

holotype).
Primary thallus 0.5-1 x 1.5-2 mm, soon disappearing. Podetia (2—)3-7 cm tall,
0.7-1.5 mm thick, pale to dark brown, medulla hardly melanotic at base, erect,
dichotomously, trichotomously or irregularly richly branched, axils open, at
least some podetia stout, sterile podetia have thin, subulate tips, fertile podetia
becoming thicker, forming characteristic lateral fissures and perforations,
branches often flattened. Surface highly discontinuously corticated, especially
towards the base, in part sparsely microsquamulose or with larger squamules,
sometimes verruculose, apical parts hardly pruinose. Podetial wall 120-220 um
thick, with indistinct cortex 10-20 um, medulla 150-200 um, and stereome
50-60 um, hard. Pycnidia terminal or sometimes lateral on podetia, rarely
on basal squamules, containing hyaline slime, 1.5 mm in diam, conidia 6-7
x 1 um. Apothecia 0.8-1 mm wide, terminal, on every tip on fertile podetia,
sometimes deformed, brown, spores not observed.

CHEMISTRY—K+ yellow, PD+ red, rarely PD-. CHEMOTYPE 1: Atranorin
(major), fumarprotocetraric acid (major), protocetraric acid (minor),
confumarprotocetraric acid (minor), rarely rangiformic and norrangiformic
acids, occasionally 1-4 unknown minor substances and 1-3 terpenoids.
CHEMOTYPE 2: Atranorin only, the fumarprotocetraric acid complex lacking
(then PD-) (Stenroos 1988). If chemotype 2 proves to represent a distinct taxon,
the name Cladonia novoguineensis is applicable for that (no fresh material for
DNA analysis was available).

HasitatT— Terrestrial in bogs, grasslands, road banks, rarely on tree stumps,
at 1500-4400 m in Papua New Guinea (Stenroos 1988: 126).

DISTRIBUTION—Australia (New South Wales, Queensland), Indonesia
(Java), New Caledonia, New Zealand (North Island), Papua New Guinea,
Philippines (Luzon).

98 ... Ahti, Pino-Bodas, & Stenroos

ADDITIONAL SPECIMENS EXAMINED (See also Stenroos 1988) — NEW CALEDONIA,
between Yaté and Goro, 1966, Hill 11741 (BM, H). PAPUA NEW GUINEA, EAsTERN
HIGHLANDS, Mt. Wilhelm, Pindaunde Lake, 3540 m, 1965, A. C. Jermy 5327 (BM, H;
richly fertile!)}; MOROBE PRov., Mt. Sarawaket Southern Range, 4 km NEE of Lake
Gwam, 2850 m, 1981, T. Koponen 31662 (H, LAE); SOUTHERN HIGHLANDS, Mt. Giluwe,
3500 m, 1982, H. Streimann 24151 (CANB, H, LAE; chemotype 2). PHILIPPINES,
Luzon, Mountain Prov., Alab on road Baguio to Bontoc, 850 m, 1976, W. S. Gruezo
2030 (CAHP, H).

Cladonia pseudofissa (Asahina) Ahti, Pino-Bodas & S. Stenroos, stat. et
comb. nov. Fic. 3B
MycoBank MB 809612
= Cladonia rangiformis var. pseudofissa Asahina, Fl. E. Himalaya: 595. 1966.
TyPE: India, Sikkim, Jongri, 4000 m, 1960, M. Togashi 157 (TNS, holotype).
= Cladonia erythrosperma var. thomsonii Vain., Acta Soc. Fauna FI.
Fenn. 4: 376. 1887, syn. nov. [non Cladonia thomsonii Ahti].
Type: India, "India orientalis’, [1847-51], J. D. Hooker & T. Thomson 2145 (PC,
holotype).
= Cladonia rangiformis var. incurva Mull. Arg., Flora 74: 72.
1891, syn. nov. [non Cladonia incurva Ahti].
Type: India, Himachal Pradesh (?), Damodar [“Damdar”] Valley, 4200 m, J. F Duthie
(BM, holotype).

The earliest names for C. pseudofissa are C. erythrosperma var. thomsonii and C.
rangiformis var. incurva, but they cannot be adopted at species level because these
epithets are already occupied in Cladonia by C. thomsonii Ahti (1978) and C. incurva
Ahti (1961).

Primary thallus rarely visible, soon disappearing, consisting of minute
squamules. Podetia 1-4(-5) cm tall, to 1 mm thick, almost white, ashy grey
or more rarely dark brown, medulla clearly melanotic at base; often very
delicate, ascendant to erect, usually somewhat curly in appearance, without any
stouter sterile podetia, repeatedly subcorymbosely dichotomously branched,
apices subulate, ascyphose, axils usually perforated. Fertile podetia somewhat
thicker. Surface very matt, minutely fibrous or pruinose, often very smooth and
almost continuously corticate, with a few very small squamules, longitudinally
somewhat fissured, esorediate. Podetial wall 150-160 um thick, with distinct
cortex 10-15 um, medulla 75-100, stereome 25-50 um thick, softish rather
than cartilaginous. Pycnidia always at tips of podetia, shortly stalked, pyriform,
black, c. 2 x 1 mm; conidia or colour of slime not observed. Apothecia dark
brown, 0.5-1 mm, subglobose, on tips of flattened branchlets, spores not
observed.

CHEMISTRY—K+ yellow, PD+ orange red. Contains atranorin and the
fumarprotocetraric acid complex.

HasitatT—On ground, often on thin soil over rocks.

Cladonia pseudofissa comb. nov. (Southeast Asia) ... 99

Fic. 3. A: Cladonia corymbescens (Papua New Guinea, T. Koponen 31662, H).
B: Cladonia pseudofissa (Nepal, P. Ozenda 147, H). Scale bar = 1 cm.

DISTRIBUTION—Bhutan, China (Sichuan, Tibet, Yunnan), India (Arunachal
Pradesh, Himachal Pradesh, Kerala, Sikkim, Tamil Nadu, Uttarkhand, West
Bengal), Malaysia (Sabah), Nepal, Thailand.

ADDITIONAL SPECIMENS EXAMINED — BHUTAN, Paro District: below Jaley la
Dzong, 2500-3000 m, 1998, U. Sochting 9206 (C, H; DNA voucher, Stenroos et al. (2002)

100 ... Ahti, Pino-Bodas, & Stenroos

and GenBank, as C. corymbescens). CHINA, SICHUAN: Miyi Co., 3200 m, 1983, L. S.
Wang 83-834 (H, KUN). TrpeT: Kang Ding Co., 1976, Z. Mu 5304 (H, KUN); YUNNAN:
Lijiang Co., 1985, L. S$. Wang 85-140 (H, KUN). INDIA, Srxxim: North Sikkim
District, Phune-Yakche, 3000 m, 1996, G. P. Sinha 1103 (ASSAM, H). UTTARAKHAND:
Pithoragarh District, Satgarh-Dhawj, 2700 m, 1989, D. K. Upreti L-18438 (H, LWG).
[For details of further Indian collections, see Rai et al. 2014, as C. corymbescens.] NEPAL,
FARWESTERN REGION: Seti, Dadachaur, Nhuna Khola, 2700 m, 1973, P. Ozenda 147C
(Gy Eh).

Discussion

Cladonia pseudofissa was included by Abbayes (1974) in C. corymbescens.
He also thought that C. perfossa Nuno, described from Taiwan, belongs
here. However, as discussed by Ahti & Lai (1979) and Stenroos (1988: 126),
C. perfossa is probably a distinct species (podetia short and thick; a topotype
re-examined in H), although still very poorly known.

Asahina (1966) clearly included the present species in C. rangiformis because
he found that it contains atranorin in addition to fumarprotocetraric acid;
such a strain is known in C. rangiformis. In habit C. pseudofissa does resemble
C. rangiformis to some extent, but C. rangiformis is not known to range east of
northwestern Iran (Litterski & Ahti 2004: 233). They are distinguished by the
longitudinally split podetia and melanotic part at the base of C. pseudofissa.

The two segregated species generally differ in habit and colour,
C. corymbescens being taller, erect, brownish (much resembling C. furcata
(Huds.) Schrad.) while C. pseudofissa is smaller, thinner, not readily erect,
and with an ashy grey or whitish tint. In addition, C. corymbescens is hardly
blackening (melanotic) at base, whereas C. pseudofissa is clearly blackening.

The phylogenetic analyses by Stenroos et al. (2002) demonstrated that
C. corymbescens was not closely related to the species of the C. furcata group
(C. farinacea (Vain.) A. Evans, C. furcata, C. multiformis G. Merr., and
C. scabriuscula (Delise) Nyl.); it seemed to be related to C. petrophila R.C.
Harris and C. apodocarpa Robbins instead. The phylogenetic study of Cladonia
in Thailand by Parnmen et al. (2008) grouped one specimen corresponding to
C. pseudofissa with C. rudis Ahti & Parnmen. Morphologically this species
is closely related to the species of C. furcata (Ahti et al. 2008). Our present
phylogenetic analyses do not allow conclusions on the phylogenetic
relationships of C. corymbescens and C. pseudofissa. Until further Cladonia
species are included in the phylogenetic analyses, the placement of these species
will remain uncertain. The ML and Bayesian analyses suggest that they might be
related, but this relationship is not supported, and the branches in both analyses
are very long. The MP analysis groups Cladonia pseudofissa with C. ceratophylla
and C. pityrophylla, but this relationship is not well supported either. A denser
sampling of taxa is needed to clarify the phylogenetic relationships between
the species.

Cladonia pseudofissa comb. nov. (Southeast Asia) ... 101

As far as is known, the geographic ranges of C. corymbescens and C. pseudofissa
do not overlap; C. corymbescens is primarily Australasian to Melanesian and
C. pseudofissa is Himalayan in distribution. However, the Cladonia flora of
Indonesia is very poorly known.

Various data support C. pseudofissa asa species distinct from C. corymbescens.
The phylogenetical analyses based on two independent loci separate Cladonia
pseudofissa and C. corymbescens into two well-supported clades, clades that
are also supported in the single gene analyses (data not shown). According to
the genealogical concordance in phylogenetic species recognition (Taylor et al.
2000), these clades represent different species. In addition, morphological and
distributional differences confirm that they are two different taxa.

Acknowledgments

We are grateful for fresh material of Cladonia collected by Dr. Maarten Christenhusz
(London) in New Caledonia. We are also grateful to Dr. Philippe Clerc (Geneva), who
sent our herbarium (H) a large collection of Cladonia from the Himalayas. Dr. Jaana
Haapala helped us in photographing. Our sincere thanks to Dr. A. R. Burgaz and
Dr. M. Piercey-Normore for their valuable comments to improve the manuscript.

Literature cited

Abbayes H des. 1974. Cladonia du Népal. Lichenes, Cladoniaceae (Flechten des Himalaya 9).
Khumbu Himal 6(2): 111-116. Innsbruck, Miinchen: Universitatsverlag Wagner.

Ahti T. 1961. Taxonomic studies on reindeer lichens (Cladonia, subgenus Cladina). Ann. Bot. Soc.
Zool. Bot. Fenn. “Vanamo’ 32(1): 1-160.

Ahti T. 1978. Two new species of Cladonia from western North America. Bryologist 81: 334-338.

Ahti T. 1993. Names in current use in the Cladoniaceae (lichen-forming ascomycetes) in the ranks
of genus to variety. Regnum Vegetabile 78: 59-106.

Ahti T, Lai MJ. 1979. The lichen genera Cladonia, Cladina and Cladia in Taiwan. Ann. Bot. Fenn.
16: 228-236.

Ahti T, Parnmen S, Mongkolsuk P. 2008. Three new species of Cladonia from Thailand. Sauteria
i Regal Ese be

Argitello A, Del Prado R, Cubas P, Crespo A. 2007. Parmelia quercina (Parmeliaceae, Lecanorales)
includes four phylogenetically supported morphospecies. Biol. J. Linn. Soc. 91: 455-467.
http://dx.doi.org/10.1111/j.1095-8312.2007.00810.x

Asahina Y. 1966. Lichens. 592-610, in: H Hara (ed.). The flora of Eastern Himalaya. University of
Tokyo.

Awasthi DD, Ahti T. 2007. Cladonia P. Browne (Cladoniaceae). 79-116, in: DD Awasthi (ed.).
A compendium of the macrolichens from India, Nepal and Sri Lanka. Dehra Dun: Bishen
Singh Mahendra Pal Singh.

Fontaine KM, Ahti T, Piercey-Normore MD. 2010. Convergent evolution in Cladonia gracilis and
allies. Lichenologist 42: 323-338. http://dx.doi.org/10.1017/S0024282909990569

Gardes M, Bruns TD. 1993. ITS primers with enhanced specificity for Basidiomycetes.
Application to the identification of mycorrhizae and rusts. Molecular Ecology 2: 113-118.
http://dx.doi.org/10.1111/j.1365-294X.1993.tb00005.x

Hall TA. 1999. BioEdit: a user-friendly biological sequence alignment editor and analysis program
for Windows 95/98/NT. Nucleic Acids Symp. Ser. 41: 95-98.

102 ... Ahti, Pino-Bodas, & Stenroos

Huelsenbeck JP, Ronquist F. 2001. MRBAYES: Bayesian inference of phylogenetic trees.
Bioinformatics 17: 754-755. http://dx.doi.org/10.1093/bioinformatics/17.8.754

Kelly LJ, Hollingsworth PM, Coppins BJ. 2011. DNA barcoding of lichenized fungi demonstrated
high identification success in a floristic context. New Phytol. 191: 288-300.
http://dx.doi.org/10.1111/j.1469-8137.2011.03677.x

Kotelko R, Piercey-Normore MD. 2010. Cladonia pyxidata and C. pocillum; genetic evidence to
regard them as conspecific. Mycologia 102: 534-545. http://dx.doi.org/10.3852/09-030

Litterski B, Ahti T. 2004. World distribution of selected European Cladonia species. Symb. Bot.
Ups. 34(1): 205-236.

Lutzoni E, Kauff F, Cox C, McLaughlin D, Celio G, Dentinger B, Padamsee M, Hibbett D, James
TY, Baloch E, Grube M, Reeb V, Hofstetter V, Schoch C, Arnold AE, Miadlikowska J, Spatafora
J, Johnson D, Hambleton S, Crockett M, Shoemaker R, Sung G-H, Liicking R, Lumbsch T,
O’Donnell K, Binder M, Diederich P, Ertz D, Gueidan C, Hansen K, Harris RC, Hosaka K,
Lim Y-W, Matheny B, Nishida H, Pfister D, Rogers J, Rossman A, Schmitt I, Sipman H, Stone J,
Sugiyama J, Yahr R, Vilgalys R. 2004. Assembling the fungal tree of life: progress, classification,
and evolution of subcellular traits. Am. J. Bot. 91: 1446-1480.
http://dx.doi.org/10.3732/ajb.91.10.1446

Muggia L, Pérez-Ortega S, Fryday A, Spribille T, Grube M. 2014. Global assessment of genetic
variation and phenotypic plasticity in the lichen-forming species Tephromela atra. Fungal
Diversity 64: 233-251. http://dx.doi.org/10.1007/s13225-013-0271-4

Nufez-Zapata J, Divakar PK, Del-Prado R, Cubas P, Hawksworth DL, Crespo A. 2010. Conundrums
in species concepts: the discovery of a new cryptic species segregated from Parmelina tiliacea
(Ascomycota: Parmeliaceae). Lichenologist 43: 603-616.
http://dx.doi.org/10.1017/S002428291100051X

Nylander JAA. 2004. MrModelTest 2.1. Program distributed by the author. Uppsala: Evolutionary
Biology Centre, Uppsala University.

Nylander JAA, Wilgenbusch DL, Warren, JC, Swofford DL. 2008. AWTY (Are we there yet?):
A system for graphical exploration of MCMC convergence in Bayesian phylogenetics.
Bioinformatics 24: 581-583. http://dx.doi.org/10.1093/bioinformatics/btm388

Orange A, James PW, White FJ. 2001. Microchemical methods for the identification of lichens.
British Lichen Society, London.

Parnmen S, Rangsiruji A, Mongkolsuk P, Ahti T. 2008. Phylogenetics of lichens in the genus
Cladonia (Cladoniaceae) in northern and northeastern Thailand. Sauteria 15: 385-402.

Parnmen S, Leavitt SD, Rangsiruji A, Lumbsch HT. 2013. Identification of species in the Cladia
aggregata group using DNA barcoding (Ascomycota: Lecanorales). Phytotaxa 115: 1-14.
http://dx.doi.org/10.11646/phytotaxa.115.1.1

Pino-Bodas R, Burgaz AR, Martin MP. 2010a. Elucidating the taxonomic rank of Cladonia subulata
versus C. rei (Cladoniaceae). Mycotaxon 113: 311-326. http://dx.doi.org/10.5248/113.311

Pino-Bodas R, Martin MP, Burgaz AR. 2010b. Insight into the Cladonia convoluta-C. foliacea
(Cladoniaceae, Ascomycota) complex and related species, revealed through morphological,
biochemical and phylogenetic analyses. Syst. Biodivers. 8: 575-586.
http://dx.doi.org/10.1080/14772000.2010.532834

Pino-Bodas R, Burgaz AR, Martin MP, Lumbsch HT. 2011. Phenotypical plasticity and homoplasy
complicate species delimitation in the Cladonia gracilis group (Cladoniaceae, Ascomycota).
Org. Divers. Evol. 11: 343-355. http://dx.doi.org/10.1007/s13127-011-0062-2

Pino-Bodas R, Burgaz AR, Martin MP, Lumbsch HT. 2012a. Species delimitations in
the Cladonia cariosa group (Cladoniaceae, Ascomycota). Lichenologist 44: 121-135.
http://dx.doi.org/10.1017/S002428291100065X

Cladonia pseudofissa comb. nov. (Southeast Asia) ... 103

Pino-Bodas R, Ahti T, Stenroos S, Martin MP, Burgaz AR. 2012b. Cladonia conista and C. humilis
(Cladoniaceae) are different species. Bibl. Lichenol. 108: 161-176.
http://dx.doi.org/1436-1698/2012/108-161

Pino-Bodas R, Martin MP, Burgaz AR. 2012c. Cladonia subturgida and C. iberica (Cladoniaceae)
form a single, morphologically and chemically polymorphic species. Mycol. Prog. 11: 269-278.
http://dx.doi.org/10.1007/s11557-011-0746-1

Pino-Bodas R, Ahti T, Stenroos S, Martin MP, Burgaz AR. 2013a. Multilocus approach to species
recognition in the Cladonia humilis complex (Cladoniaceae, Ascomycota). Am. J. Bot. 100:
664-678. http://dx.doi.org/10.3732/ajb.1200162

Pino-Bodas R, Martin MP, Burgaz AR, Lumbsch HT. 2013b. Species delimitation in Cladonia
(Ascomycota): a challenge to the DNA barcoding philosophy. Mol. Ecol. Resour. 13: 1058-1068.
http://dx.doi.org/10.1111/1755-0998.12086

Rai H, Khare R, Upreti DK, Ahti T. 2014. Terricolous lichens of India: taxonomic keys and
description. 17-294, in: H Rai, DK Upreti (eds). Terricolous lichens in India 2. Springer,
Heidelberg. http://dx.doi.org/10.1007/978-1-4939-0360-3_2

Schoch CL, Seifert KA, Huhndorf S, Robert V, Spouge JL, Levesque A, Chen V, Fungal Barcoding
Consortium. 2012. Nuclear ribosomal internal transcribed spacer (ITS) region as a universal
DNA barcode for Fungi. Proc. Natl. Acad. Sci. USA 109: 6241-6246.
http://dx.doi.org/10.1073/ pnas.1117018109.

Stamatakis A. 2006. RAxML-VI-HPC: maximum likelihood-based phylogenetic analyses with
thousands of taxa and mixed models. Bioinformatics 22: 2688-2690.
http://dx.doi.org/10.1093/bioinformatics/btl446

Steinova J, Stenroos S, Grube M, Skaloud P. 2013. Genetic diversity and species delimitation of
the zeorin-containing red-fruited Cladonia species (lichenized Ascomycota) assessed with ITS
rDNA and £-tubulin data. Lichenologist 45: 665-684.
http://dx.doi.org/10.1017/S0024282913000297

Stenroos S. 1988. The family Cladoniaceae in Melanesia. 3. Cladonia sections Helopodium, Perviae
and Cladonia. Ann. Bot. Fenn. 25: 117-148.

Stenroos S, Hyvonen J, Myllys, L, Thell, A, Ahti T. 2002. Phylogeny of the genus Cladonia s. lat.
(Cladoniaceae, Ascomycetes) inferred from molecular, morphological, and chemical data.
Cladistics 18: 237-278. http://dx.doi.org/10.1006/clad.2002.0202

Swofford DL. 2002. PAUP*: phylogenetic analysis using parsimony (*and other methods), version
4.0b10. Sinauer, Sunderland, Massachusetts, USA.

Taylor JW, Jacobson DJ, Kroken S, Kasuga T, Geiser DM, Hibbett DS, Fisher MC. 2000.
Species recognition and species concepts in fungi. Fungal Genet. Biol. 29: 21-32.
http://dx.doi.org/10.1006/fgbi.2000.1228

Vainio EA. 1887. Monographia Cladoniarum universalis 1. Acta Soc. Fauna et Flora Fenn. 4: 1-509.

Yahr R, Vilgalys R, DePriest PT. 2006. Geographic variation in algal partners of Cladonia subtenuis
(Cladoniaceae) highlights the dynamic nature of a lichen symbiosis. New Phytol. 171: 847-860.
http://dx.doi.org/10.1111/j.1469-8137.2006.01792.x

MY COTAXON

http://dx.doi.org/10.5248/130.105
Volume 130, pp. 105-130 January-March 2015

Morphology and phylogeny of four new Lactarius species from
Himalayan India

KANAD Das", ANNEMIEKE VERBEKEN’, & JORINDE NUYTINCK”?

"Botanical Survey of India, Cryptogamy Unit, Central National Herbarium,
Howrah 711103, W.B., India

?Ghent University, Department of Biology, Research Group Mycology,
K.L. Ledeganckstraat 35, BE 9000, Gent, Belgium

*Naturalis Biodiversity Center, Section National Herbarium of the Netherlands,
PO. Box 9517, 2300RA Leiden, The Netherlands

* CORRESPONDENCE TO: daskanadbsi@gmail.com

ABSTRACT —Four new species of Lactarius are described from Himalayan India. Lactarius
olivaceoglutinus, L. pyriodorus, and L. yumthangensis belong to L. subg. Piperites and
L. indochrysorrheus is closely related to some representatives of L. subg. Russularia. An ITS
based phylogeny confirms the phylogenetic placement of the four new species, although the
monophyly of neither Lactarius subg. Piperites nor L. subg. Russularia can be confirmed.
The ITS data also suggest that the Indian species are closely related to some European and
American species.

Key worps — ectomycorrhizal fungi, macrofungi, Russulaceae, Sikkim

Introduction

After segregation of the well-known ectomycorrhizal milkcaps into Lactarius
Pers. and Lactifluus (Pers.) Roussel (Buyck et al. 2008, 2010), Lactarius sensu
novo comprises three subgenera (Verbeken & Nuytinck 2013): L. subg. Piperites
(Fr. ex J. Kickx f.) Kauffman, L. subg. Russularia (Fr. ex Burl.) Kauffman, and
L. subg. Plinthogali (Burl.) Hesler & A.H. Sm. Lactarius subg. Piperites can
be characterized as follows: pileus sticky to slimy/glutinous (more rarely dry
and shiny), often hairy and/or zonate or with watery spots; stipe dry or sticky,
often scrobiculate (Heilmann-Clausen et al. 1998, Basso 1999). In contrast,
representatives of L. subg. Russularia typically have dry (rarely somewhat
sticky) caps and stipes and colours that are predominantly orange or various
tinges of brown (Heilmann-Clausen et al. 1998).

106 ... Das, Verbeken, & Nuytinck

Sikkim, a small (0.22% of the land surface) state in India, lies in Eastern
Himalaya, which is part of the Himalaya Hotspot, one of the 34 Global
Biodiversity Hotspots (www.biodiversityhotspots.org). Within a very small
geographical area it is substantially diverse in flora, fauna, and mycobiota, the
latter seriously underexplored. During mycological expeditions to different
temperate to subalpine areas of the North District of Sikkim in 2009 (KD &
AV), 2011 (KD), and 2012 (KD), we collected ectomycorrhizal (ECM) fungi,
with Lactarius s.l. one of the dominant ECM genera in Dombang, Shingba
Rhododendron Sanctuary, and Zema. Dombang is a subalpine coniferous to
mixed (coniferous & broadleaf) forest dominated by Picea spinulosa, Abies
densa, Tsuga dumosa, and Larix griffithii. Shingba Rhododendron Sanctuary
contains subalpine coniferous and mixed forests distributed in Yumthang
valley and its adjoining areas; apart from several Rhododendron species, this
protected area is dominated by Abies densa, Picea spinulosa, Tsuga dumosa,
Larix griffithii, Magnolia globosa, M. campbellii, Acer pectinatum, and Betula
utilis. Zema is a subalpine coniferous forest dominated by Abies densa.

Although Sikkim harbours a large number of russulaceous taxa, only 32
species (out of ca 210 taxa recorded from India) have been reported (Berkeley
1852; Das 2009; Das et al. 2010, 2013; Das & Verbeken 2011, 2012; Van de Putte
et al. 2012). We describe here three new species of Lactarius subg. Piperites
(L. olivaceoglutinus, L. pyriodorus, and L. yumthangensis) and one new species
of L. subg. Russularia (L. indochrysorrheus).

Materials & methods

Morphological study

Macromorphological characters were described from the fresh basidiomata in
daylight. Colour codes and terms follow the COLOUR IDENTIFICATION CHART OF
BRITISH FuNGus FLora (Henderson et al. 1969, here prefixed by “a:”) or the METHUEN
HANDBOOK OF CoLourR (Kornerup & Wanscher 1981, here prefixed by “b:”). Spore
print colour codes follow Kranzlin (2005, here prefixed by “c:”).

Micromorphological structures such as basidia, hymenial cystidia, pileipellis,
stipitipellis, etc. were observed from free-hand sections of dry samples mounted in
a mixture of 5% KOH, 30% Glycerol, Phloxine, and Cotton Blue using an Olympus
CX41 compound microscope. Spores and spore ornamentation were studied in Melzer’s
reagent. Drawings were made from SEM images obtained in different magnifications.
Spore measurements were calculated based on 20 basidiospores per specimen;
dimensions represent minimum-mean-maximum length x minimum-mean-
maximum width, and Q = length/width ratio. Herbarium names are after Holmgren et

al. (1990).

DNA extraction, PCR amplification and sequencing
DNA was extracted from dried fruiting bodies according to Nuytinck & Verbeken
(2003) with slight modifications (Van de Putte et al. 2010). The internal transcribed

Lactarius spp. nov. (India) ... 107

TABLE 1. Collections of Lactarius and allied species used for molecular analyses.

SPECIES

Lactarius akahatsu

L. albocarneus

alboscrobiculatus
aspideus
atroviridis
auriolla
indochrysorrheus
azonites

Reis ia fot Enea

brunneoviolaceus

caespitosus
camphoratus
chichuensis

Ea itech

chrysorrheus

L. aff. chrysorrheus

L. citriolens

L. controversus
L. crassiusculus
L. cyathuliformis
L. flexuosus

L. formosus
L. fuliginosus
L. helvus

L. lilacinus
L. luridus

L. montoyae

L. necator

L. olivaceoglutinus
L. pallescens

L. peckii

L. pseudouvidus

VOUCHER

Verbeken 04-141
Verbeken 98-071
Verbeken 98-080

Le 175

Walleyn 3815
Verbeken 05-306
Walleyn 1601

KD 11-002 (Holotypus)
Verbeken 00-124
Verbeken 04-220
Verbeken 04-249
Walleyn 1605
Eberhardt 24.08. 04-8

Oberwinkler 46773
Wang 1236

Nuytinck 01-089
D’Hooge 08-020
Eberhardt 04.10. 02-8
Verbeken 04-212
Verbeken 05-359
Eberhardt 20.09.04-3
Verbeken 00-117

Le 369

Eberhardt 04.09.04-3
Walleyn 2136
Walleyn 3178
Eberhardt 06.09.02-1
Le 382 (Holotypus)
Basso 97-24
Eberhardt 08.09. 04-1
Walleyn 3774

Taylor 2003066
Eberhardt 17.09.04-3
Eberhardt 10.10. 04-4
Berteloot 11-011
Berteloot 11-012
Taylor 2004254
Walleyn 1455

KD 1065 (Holotypus)
Verbeken 04-231

KD 11-103 (Holotypus)

Nuytinck 04-020
Eberhardt 24.08. 04-14
Eberhardt 24.08. 04-13

HERBARIUM
GENT

GENT

CMU, SFSU, GENT
GENT

GENT

BSHC, GENT
GENT

GENT

UPS

TUB
HKAS
GENT
GENT
UPS
GENT
GENT
UPS
GENT

CMU, SFSU

UPS

GENT

URS

CMU, SFSU, GENT
priv. herb.

UPS

GENT

UPS

GENT

UPS
GENT
BSD
GENT

BSHC, GENT

GENT
UPS
UPS

ORIGIN

Thailand
France

France
Thailand

Czech Rep.

USA
Sweden
India
Belgium
France
France
Sweden
Sweden

Germany
China
Belgium
France
Italy
USA
USA
Sweden
Italy
Thailand
Sweden
Sweden

Czech Rep.

Sweden
Thailand
Sweden
Sweden
Belgium
Sweden
Sweden
Sweden
Belgium
Belgium
Sweden
Belgium
India
France
India

USA
Sweden
Sweden

GENBANK

KF133269
KJ742389
KF241545
EF141538
KJ742390
KF133270
KF133257
KJ742391
KF241540
KJ742392
KJ742393
KJ742394
KJ742395
FJ845421
AY606945
KF475766
KJ742396
KJ742397
KF133261
KJ742398
KJ742399
DQ422003
KF241544
EF560684
KF133266
KJ742400
KJ742401
DQ421992
EF141549
JQ446111
KF133263
KF133275
KJ742402
KJ742403
KJ742404
KF241547
KJ742405
KJ742406
KJ742407
EF560673
KF133276
KJ742408
DQ974747
KF133277
KJ742409
KJ742410

108 ... Das, Verbeken, & Nuytinck

L. pubescens Eberhardt 15.09.02-2 UPS Sweden DQ421996
L. pyriodorus KD 11-027 (Holotypus) | BSHC, GENT India KJ742411
L. quieticolor Eberhardt 10.09.04-1 UPS Sweden DQ422002
L. quietus Eberhardt 16.09.04-6 UPS Sweden KF133264
L. rufus Nuytinck 02-008 GENT Norway KF241543
L. sphagneti Walter 083 TUB Germany KJ742412
L. subdulcis Vervisch 06-024 GENT Belgium KF133279
L. subplinthogalus Verbeken 04-219 GENT USA KF241539
L. subsericatus Eberhardt 11.10.04-8 UPS Sweden DQ422011
L. thyinos Voitk 23-08-04 priv. herb. Canada KF133271
L. torminosus Walleyn 3183 GENT Czech Rep. KF133281
L. trivialis Van de Putte 10-011 GENT Russia KJ742413
Walleyn 3179 GENT Czech Rep. KJ742414
Eberhardt27.08. 02-17a UPS Sweden DQ421991
L. uvidus Van de Putte 10-027 GENT Russia KF241546
Nuytinck 01-033 GENT Finland KJ742415
Walleyn 1237 GENT France KJ742388
Walleyn 2119 GENT Sweden KJ742416
Eberhardt 28.08. 02-24 UPS Sweden KJ742417
L. vietus Eberhardt 11.10.04-1 UPS Sweden KF133267
L. vinaceorufescens Nuytinck 07-018 GENT Canada KF241542
L. yumthangensis KD 11-147 (Holotypus) | BSHC, GENT India KJ742418
Lactifluus volemus Eberhardt 09.08. 04-5 UPS Sweden DQ422008
Lactifluus vellereus Eberhardt 20.09.04-22 UPS Sweden DQ422034
Gace x Buyck 02-107 PC USA DQ421984
M. zonaria Desjardin 7442 SFSU, PC, BBH Thailand DQ421990
Russula cyanoxantha Eberhardt 29.09. 02-2 UPS France DQ422033
R. nigricans Eberhardt 20.09.04-7 UPS Sweden DQ422010

spacer region of the nuclear ribosomal DNA (ITS) was amplified and sequenced using
primers ITS1-F and ITS4 (White et al. 1990, Gardes & Bruns 1993). PCR amplification
protocols follow Le et al. (2007); sequencing was conducted with an ABI 3730XL or
ABI 3700 by MacroGEN (Amsterdam, The Netherlands). Sequences were assembled
and edited with Sequencher™ v4.9 (GeneCodes Corporation, Ann Arbor, Michigan,
US.A.). Specimens and sequences used in the phylogenetic analysis are shown in
TABLE l.

Alignment and phylogenetic analyses

Alignment was conducted with the on-line version of MAFFTv7 (Katoh & Standley
2013), applying the E-INS-I strategy, a very slow method recommended for fewer than
200 sequences with multiple conserved domains and long gaps. The alignment was
manually refined in BioEdit v7.0.9.0 (Hall 1999). Ambiguously aligned positions were
detected using Gblocks v0.91b (Castresana 2000), specifying less stringent conditions
than default. The minimum number of sequences for a conserved position and for a
flank position was set to half the number of sequences, the number of contiguous non-
conserved positions was set to 10, and the minimum length of a block after gap cleaning
to 5 and positions with gaps were not treated differently from other positions. The ITS

Lactarius spp. nov. (India) ... 109

L. pallescens DQ874747 USA
L. brunneoviolaceus AV04,220 FRA
L. brunneoviolaceus AV04,249 FRA
L. brunneoviolaceus RW1605 SWE
L. brunneoviolaceus UE24.08.04.8 SWE
se L. luridus AT2003066 SWE
L. luridus UE17.09.04.3 SWE
SrA |e. luridus UE10.10.04.4 SWE
L, luridus OB11.012 BEL
88 L. luridus OB11.011 BEL
L. luridus AT2004254 SWE
98] [IL iuridus RW1455 BEL
L. pyriodorus KD11.027 IND
L. aspideus RW3815 CZE
L. pseudouvidus UE24.08.04.14 SWE
egbL. pseudouvidus UE24.08.04.13 SWE
L. uvidus RW1237 FRA
L. wvidus RW2119 SWE
7 L. uvidus JNO1.033 FIN
92) L. uvidus UE28,08,02.24 SWE
L. wvidus KVP 10.027 RUS
L formosus LTH382 THA
L. olivaceoglutinus KD11.103 IND
L. caespitosus FJ845421
L. albocameus AV98.071 FRA
964 L. albocarneus AV98.080 FRA
L. alboscrobiculatus LTH175 THA
64 L, torminosus RW3183 CZE
L. pubescens UE15.09.02.2 SWE
L.controversus AV00.117 ITA
Ey L.quieticolor UE10.09.04.1 SWE
L. akahatsu JNO4.141 THA
84 L. thyinos AV23.08.04 USA
L. auriolla RW1601 SWE
0b L. citriolens UE20.09.04.3 SWE

L quietus UE 16.09.04.6 SWE
58 L. helws UE08.09.04.1 SWE
L. chichuensis XW1236 CHI
81 L. camphoratus FO46773 GER
L. cyathuliformis UE04.09.04.3 SWE
L, subdulcis JV06.024 BEL
3 L. subsericatus UE11.10.04.8 SWE

86 96

7 L. sphagneti LW083 GER
8 L. rufus JNO2,008 NOR
77 86) L. chrysonheus JNO1.089 BEL
100 fl. chrysorheus ED08.020 FRA
73 Lchrysortheus UE04.10.02.8 ITA
= 100 L. aff. chrysormheus AVO4.212 USA
L. aff. chrysomheus AVO5.359 USA
63|7—L. indochrysorrheus KD11.002 IND
64 L. vinaceorufescens JNO7.018 CAN
L. peckii JNO4.020 USA
L. yumthangensis KD11.147 IND
L, fexuosus RW2136 SWE
L flexuosus RW3178 CZE
SIL. flexuosus UE0S.09.02.1 SWE
L. trivalis UE27.08.02-17a SWE
L. trivalis KVP10.010 RUS
L. trivialis RW3179 CZE

100!
L lilacinus RW3774 BEL
L. atroviridis AVO5.306 USA
L. necator AV04.231 FRA
L. vetus UE11.10.04.1 SWE
100 ‘L. azonites AVO0.124 BEL
97) L. fuliginosus MTB97-24 SWE
L. subplinthogalus AV04,219 USA

77] 76 L. montoyae typus IND
L. crassiusculus LTH369 THA
100, 'M. ochricompacta BB02.107 USA
M. zonaria DED7442 THA
R. cyanoxantha UE29.09.02.2 SWE
cy R nigricans UE20.09.04.7 SWE
LE. volemus 09.08.04.5 SWE
% Lt vellereus UE20.09.04-22 SWE

“1
0.05

PLATE 1. The obtained ML topology based on ITS sequences of Lactarius, Multifurca, Russula, and
Lactifluus species. Bootstrap values >50% are indicated. Names in bold are the new Indian species
described in this paper. The scale bar represents the number of nucleotide changes per site.

sequences were partitioned into 5 partitions: the ribosomal genes 18S, 5.88, and LSU
and the spacer regions ITS1 and ITS2.

Maximum Likelihood (ML) analysis was performed in RAXML v7.0.3 (Stamatakis
2006), combining a ML search with the Rapid Bootstrapping algorithm for 1000
replicates. The model GTRGAMMA was estimated for each partition separately.

Results

Phylogeny

PLaTE 1 shows the obtained ML topology with bootstrap support (BS)
values >50% displayed. The tree shows a well-supported genus Lactarius (97%
BS). Within Lactarius, L. subg. Plinthogali receives a high support value (100%
BS), but monophyly of neither Lactarius subg. Piperites nor L. subg. Russularia
(sensu Heilmann-Clausen et al. 1998) is supported. Lactarius pyriodorus,
L. olivaceoglutinus, and L. yumthangensis are closely related to species

110 ... Das, Verbeken, & Nuytinck

traditionally placed in L. subg. Piperites. Lactarius indochrysorrheus is closely
related to the European L. chrysorrheus Fr. and the North American L. vinaceo-
rufescens A.H. Sm. and L. aff. chrysorrheus, which are assigned to L. subg.
Russularia in morphology-based taxonomy.

Species delimitation for this paper is mainly based on morphological
features. We were not able to include enough specimens/sequences from the
newly proposed species to test species delimitation in the phylogenetic tree.

Taxonomy

Lactarius indochrysorrheus K. Das & Verbeken, sp. nov. PLATES 2, 3, 6A,B,E
MycoBank MB804887

Differs from Lactarius chrysorrheus by its more viscid pileus with its pileipellis an
ixocutis covered by a thick gelatinous layer.

Type: India. Sikkim: North District, Dombang, 27°43’35.2”N 88°45’15.2”E, alt. 2920 m,
18.VIII.2011, K. Das, KD 11-002 (Holotype, BSHC; isotype, GENT).

EryMo_oey: an Indian look-alike of the European Lactarius chrysorrheus.

PiLEus 25-70 mm diam., convex with slightly pubescent and inrolled margin
when young, gradually planoconvex to applanate with slightly depressed centre,
rarely with a central papilla, sometimes becoming widely infundibuliform;
margin decurved with maturity, becoming very irregularly wavy; surface
smooth to greasy, viscid (sticky), shiny, brownish orange, pinkish buff to salmon
(a: 45) or orange (b: 6A6) (cinnamon (a: 10) to somewhat rusty after maturity),
gradually apricot cream to paler up to yellow (a: 5E), towards margin distinctly
zonate with several zones over the whole diameter but most dense in the center;
zones consisting of darker and watery spots; margin very faintly and shortly
striate. LAMELLAE subdecurrent, crowded (18-22/cm at pileus margin), pale
yellow (a: 4D) when young, gradually pink-spotted, finally becoming brown
to reddish brown, with lamellulae in 9 series; edge entire, concolorous. STIPE
45-65 x 6-8.5 mm, slender, subcylindrical to cylindrical or slightly widened
towards base; surface smooth, slightly greasy, strigose (hairy) at base, much
paler than the pileus, especially in the upper part (very pale pinkish), very pale
salmon to vinaceous, gradually darker up to rust to rusty tawny or pale brick
red. CONTEXT hollow in stipe, pinkish yellow (a: 4D) to pale salmon, turning
lemon yellow (a: 54) to greenish yellow (a: 57) after cut, changing to salmon
(a: 45) with FeSO, and greenish with Guaiac. LaTex abundant, white, turning
quickly to greenish yellow. Taste first mild, then becoming bitter and acrid.
Opour not distinctive. SPORE PRINT pale cream (c: 10 Y).

BASIDIOSPORES 6.4—7.4-8.5 x 5.6-6.2-6.9 um, (Q = 1.08-1.18-1.33),
subglobose to ellipsoid; ornamentation amyloid, <0.9 um high, composed
of conical to spine-shaped warts, with short or medium, irregular to regular

Lactarius spp. nov. (India) ... 111

PLATE 2: Lactarius indochrysorrheus (Holotype KD 11-002).
A. Fresh basidiomata showing lamellae and lamellulae. B. Basidiospores.
C. Marginal cells. D. Basidia. Scale bars: B = 5 um; C, D = 10 um.

112 ... Das, Verbeken, & Nuytinck

ridges which are aligned or connected and forming a partial to incomplete
reticulum; some isolated small warts present; plage sometimes distinct and
amyloid. Basrp1a 30-44 x 9-13 um, 4-spored, subclavate to ventricose;
sterigmata 2.5-4 x 1.5-2 um. PLEUROMACROCYSTIDIA 32-70 x 7.5-11 um,
fairly abundant, emergent <10-30 um, narrowly clavate, cylindric with tapered
apex or fusoid, often with mucronate to subcapitate apex, slightly thick-walled
(wall <0.5 um); content refractive. PLEUROPSEUDOCYSTIDIA filamentous, 3-5
um wide. LAMELLAR EDGE fertile with basidia, cystidia and marginal cells.
CHEILOMACROCYSTIDIA 35-55 x 8-9 um, moderately abundant, subclavate
to narrowly clavate or ventricose to fusoid sometimes with mucronate apex;
content refractive. CHEILOPSEUDOCYSTIDIA filamentous. MARGINAL CELLS
11-20 x 7-11 um, mostly clavate to subclavate, often multiseptate.
HYMENOPHORAL TRAMA with lactifers. PILEIPELLIS an ixocutis, <170 um thick,
with gelatinous layer mostly extended 10-15 um beyond the hyphal layer;
hyphae repent to suberect, <3.5 um wide, branched, septate. STIPITIPELLIS
an ixocutis, composed of repent hyphae mostly in parallel pattern; hyphae
<3.5 um wide, branched, septate. STIPE TRAMA mostly with numerous nested
sphaerocytes. CLAMP CONNECTIONS absent.

EcoLocy & DISTRIBUTION — Gregarious under Abies densa and Picea
spinulosa in subalpine coniferous or mixed (coniferous and broadleaf) forest.
August. Fairly common.

ADDITIONAL SPECIMENS EXAMINED: INDIA. SIKKIM: North District, Dombang,
27°43'35.2”N 88°45'15.2’E, alt. 2920 m, 14.VIII.2009, A. Verbeken, K. Das & K.V. Putte,
AV-KD-KVP 032 (BSHC, GENT); North District, Shingba Rhododendron sanctuary,
27°43'42.1”N 88°44'58.8”E, alt. 2889 m, 24.VIII.2011, K. Das, KD 11-082 (BSHC);
27°44’03.5”N 88°44’23.3’E, alt. 3208 m, 26.VIII.2012, K. Das, KD 11-107 (BSHC).
Notes — Zonate species with similar cap colours and milk that stains quickly
sulphur to greenish yellow are European L. chrysorrheus, North American
L. vinaceorufescens, and an undescribed American species [labeled L. aff.
chrysorrheus in the phylogenetic tree and TABLE 1]. ITS sequence comparison
suggests the new species described here is close to the three species but not
conspecific.

Lactarius chrysorrheus is less sticky, which is reflected in the pileipellis
structure, a loosely interwoven cutis without distinct slime layer (Heilmann-
Clausen et al. 1998). Lactarius vinaceorufescens differs by a pileipellis with a
relatively thin ixocutis and the lack of ixocutis in the stipitipellis (Hesler &
Smith 1979). Lactarius xanthogalus Verbeken & E. Horak, described from
Papua New Guinea (Verbeken & Horak 2000), has a zonate orange to paler cap
and latex that becomes immediately sulphur yellow but clearly differs by the
zebroid ornamented spores.

aks,

Lactarius spp. nov. (India) ...

Pleuromacrocystidia. B. Radial section through pileipellis.

PLATE 3. Lactarius indochrysorrheus (Holotype KD 11-002).
C. Pleuropseudocystidia. D. Cheilomacrocystidia. Scale bars = 10 um.

Pes

114 ... Das, Verbeken, & Nuytinck

Lactarius olivaceoglutinus K. Das & Verbeken, sp. nov. PLATES 4, 5, 6C,D,F
MycoBaAnk MB 804888
Differs from Lactarius albocarneus by its olivaceous pileus and its watery white latex that

turns faintly pinkish on bruised or cut gill tissue.

Type: India. Sikkim: North District, Shingba Rhododendron sanctuary, 27°44’19.5”N
88°44’25.9’E, alt. 3252 m, 26.VIII.2011, K. Das, KD 11-103 (Holotype, BSHC; isotype,
GENT).

EryMoLoey: named after the olive and extremely glutinous pileus of the basidiocarps.

Pireus 40-68(-120) mm diam., at first convex, gradually becoming
planoconvex with slightly depressed centre; surface smooth, viscid to highly
glutinous, olive yellow (b: 3D8) to olive brown (b: 4E8, 4F6) or golden brown
(b: 5D7) to greenish grey, grey-olive or darker towards centre and paler margin,
azonate; margin non-striate, incurved when young, decurved with maturity.
LAMELLAE adnexed to broadly adnate, crowded (17-18/cm at margin), with
lamellulae in 5-6 series, pale cream to cream; edge entire, concolorous. STIPE
40-75 x 12-17 mm, clavate to subclavate (broader towards base) or ventricose
(narrower towards apex and base); surface smooth or sometimes with shallow
depressions (but without scrobicules), sticky, pale salmon to pale yellow (b:
3A3 to 4A3), gradually whitish towards base; base strigose, pale cream to
white, sometimes with ochraceous spots. CONTEXT firm, stuffed to hollow in
stipe, cream (a: 4D) to pale cream, unchanging when exposed or with KOH,
changing to olivaceous (a: 62) to leaf green (a: 59) with Guaiac and pale green
with FeSO,. LATEX whitish, very watery, turning faintly pinkish on bruised or
cut lamellae. Taste mild and agreeable or slightly acrid. ODour indistinct.
SPORE PRINT pale yellow (c: 30Y, 2M).

BASIDIOSPORES 8.0-9.3-10.1(-11) x 6.7-7.3-7.8(-8.4) um, [Q = 1.16-1.28-
1.33(-1.44)], broadly ellipsoid to ellipsoid; ornamentation amyloid, <1.2 um
high, composed of narrow, low and high ridges aligned and parallel, forming
zebroid pattern, some connected with adjacent ones (but never forming a true
reticulum), with each high ridge bearing elongated to conical (with rounded
apex) warts; plage indistinct. Bastp1a 40-60 x 11.5-13 um, 2-4-spored,
ventricose; sterigmata 6-9 x 2-2.5 um. PLEUROMACROCYSTIDIA 57-88 xX
8-12 um, abundant, subcylindric or subclavate to fusiform with rounded,
mucronate to moniliform (rarely) apex, emergent <41 um; content dense.
PLEUROPSEUDOCYSTIDIA cylindrical to tortuous, rare, never emergent, 4-5
um wide; content refringent. LAMELLAR EDGE sterile with marginal cells
and some cystidia. CHEILOMACROCYSTIDIA 32-57 x 7-9 um, subcylindrical
to subfusiform, mostly with rounded apex. MARGINAL CELLS 13-23 x 6.5-9
um, narrowly clavate to clavate or subcapitate, slightly thick-walled, hyaline.
HYMENOPHORAL TRAMA with lactifers. PILEIPELLIS an ixotrichoderm, very
thick (often variable in thickness), 220-700 um thick, composed of erect,

Lactarius spp. nov. (India) ... 115

PLATE 4. Lactarius olivaceoglutinus (Holotype KD 11-103).
A. Fresh basidiomata showing lamellae and lamellulae. B. Basidiospores.
C. Basidia. D. Marginal cells. Scale bars: B = 5 um; C, D = 10 um.

116 ... Das, Verbeken, & Nuytinck

oe ST oy
ra

103).

Pleuromacrocystidia. B. Radial section through pileipellis.

PLATE 5. Lactarius olivaceoglutinus (Holotype KD 11-
C. Cheilomacrocystidia. D. Pleuropseudocystidia. Scale bars = 10 um.

Lactarius spp. nov. (India) ... 117

PiateE 6. Lactarius indochrysorrheus. A. Fresh basidiomata. B. Latex oozing from cut lamellae.
E. Basidiospores (SEM). Lactarius olivaceoglutinus. C, D. Fresh basidiomata. FE. Basidiospores
(SEM). Scale bars: E, F = 5 um.

branched multi-septate hyphae (<3.5 um wide) and few lactifers embedded
in a slime layer of variable thickness underlying somewhat repent hyphae.
STIPITIPELLIS an ixotrichoderm, <120 um thick, composed of erect branched

118 ... Das, Verbeken, & Nuytinck

septate hyphae (<3 um wide) embedded in a slime layer. STIPE TRAMA with
numerous nested sphaerocytes. CLAMP CONNECTIONS absent.

EcoLoGy & DiIsTRIBUTION — Gregarious under Abies densa in subalpine
coniferous or mixed (coniferous and broadleaf) forest. August-September.
Fairly common.

ADDITIONAL SPECIMENS EXAMINED: INDIA. Sikkim: North District, Shingba

Rhododendron sanctuary, 27°45’11.1”N 88°43’47.5’E, alt. 3580 m, 29.VIII.2011, K.

Das, KD 11-144 (BSHC); North District, Dombang, 27°44’53.1”N 88°44’58.8”E, alt.

3058 m, 03.IX.2011, K. Das, KD 11-191 (BSHC); 27°44’51”N 88°46'32’E, alt. 2940 m,

15.VIII.2009, A. Verbeken, K. Das & K.V. Putte, AV-KD-KVP 09-049 (BSHC, GENT).
Notes — The combination of the viscid to glutinous pileus and stipe and the
ixotrichoderm pileipellis clearly places the present species in L. sect. Pallidini.
Lactarius olivaceoglutinus can be recognized by its distinctly glutinous grey-olive
to greenish grey pileus, pale salmon to pale yellow sticky stipe, whitish watery
latex that turns the bruised lamellae faintly pinkish, large spores with mostly
zebroid patterns of ridges (that carry warts), and its occurrence under Abies
sp. ITS sequences and morphology (mainly the thick pileipellis ixotrichoderm)
suggest a close relationship with the European L. albocarneus Britzelm.
[= L. glutinopallens FH. Moller & J.E. Lange] and North American L. caespitosus
Hesler & A.H. Sm. The pileus of L. albocarneus however, is dominantly cream
to at most olivaceous buff and its latex slowly turns to sulfur-yellow (Heilmann-
Clausen et al. 1998, Kranzlin 2005) and its spore ridges never bear elongated
to conical warts. Lactarius caespitosus also has a viscid cap with similar grey to
olive brown colours, but its spores are slightly larger with a distinctly low and
non-zebroid ornamentation (Hesler & Smith 1979).

In Sikkim L. elaioviscidus K. Das & Verbeken is another species with a
glutinous olivaceous pileus and watery white latex (Das & Verbeken 2011);
however, it is distinguished from L. olivaceoglutinus by the presence of a papilla,
its greyish yellow to yellowish white stipe without pinkish tinges, latex changing
to yellow-yellowish green after long exposure, and smaller basidiospores
without zebroid ornamentation (7.0-8.1-8.7 x 6.0-6.7-7.1 tm).

Lactarius pyriodorus K. Das & Verbeken, sp. nov. PLATE 7-9, 13A,B,F
MycoBank MB 804889
Differs from Lactarius formosus by lacking bundles of glutinous hairs on the pileus.

Type: India. Sikkim: North District, Dombang, 27°44’08.7”'N, 88°45’58.3”, alt. 2975 m,
19. VIII.2011, K. Das, KD 11-027 (Holotype, BSHC; isotype, GENT).

EryMo oy: in reference to the distinct pear-like odor of the fresh basidiocarps.
PILEus 35-77 mm diam., at first convex with slightly depressed centre, gradually

becoming planoconvex with depressed centre and finally broadly to widely
infundibuliform, rarely with a small, sometimes blunt central papilla; surface

Lactarius spp. nov. (India) ... 119

Pate 7. Lactarius pyriodorus (Holotype KD 11-027).
A. Fresh basidiomata showing lamellae and lamellulae. B. Basidiospores.
C. Basidia. D. Cheilomacrocystidia. Scale bars: B = 5 um; C, D = 10 um.

120 ... Das, Verbeken, & Nuytinck

smooth to finely rugulose under the slimy surface, viscid to highly glutinous
and slimy, mostly uniformly colored but sometimes with some indistinct
fading zones, bay (a: 19) to umber (a: 18), often sienna (a: 11) towards margin
with distinct rust (a: 13) to dark brick (a: 20) or brown vinaceous (a: 25), with
broader zonations towards centre and narrower zones towards margin; margin
non-striate to very finely and shortly striate, incurved when young, decurved
with maturity, becoming very irregularly wavy to undulate. LAMELLAE broadly
adnate to decurrent, crowded (15-16/cm at margin), sometimes forked near
the stipe, with lamellulae in 5 series, whitish cream to pale cream, turning livid
vinaceous (a: 77) to lilac (a: 79) or darker violet (b: 15D4-5, 15E5) after bruising;
edge entire, concolorous. STIPE 35-72 x 10-18 mm, cylindrical or broadest in
the middle, narrower towards base, often curved and locally swollen; surface
smooth, slightly viscid or sticky, not as viscid as pileus, dingy white to pale
cream, buff (a: 52) to vinaceous buff with maturity; base usually paler, strigose.
CONTEXT firm, rather thick in pileus center and thin in outer half, very soon
hollow in stipe, white to pale cream, changing lilac (a: 77; b: 13C3-4) after
exposure, leaf green with Guaiac, unchanging with FeSO, and KOH. LaTex
white to pale cream, watery, slowly staining the lamellae or context livid
vinaceous (a: 77) to lilac (a: 79), unchanging without the contact of lamellae
or context. TasTE mild at first, becoming slightly bitter and astringent. ODOUR
fruity, very sweet, like pears (as in Inocybe corydalina Quél.). SPORE PRINT pale
cream (c: 10Y).

BASIDIOSPORES 8.0-8.9-10.5 x 6.9-7.5-8.8 um, (Q = 1.11-1.18-1.32),
subglobose to broadly ellipsoid, rarely ellipsoid; ornamentation amyloid, <1.1
um high, composed of broad ridges and isolated warts forming a partial to an
incomplete reticulum; plage inamyloid or distally amyloid. Bastp1a 45-52 x
10.5-12 um, 4-spored, rarely 2-spored, clavate to ventricose; sterigmata 6-7
x 2-2.5 um. PLEUROMACROCYSTIDIA 55-80 Xx 7.5-11 um, abundant, fusiform
with mucronate to moniliform apex, emergent <33 um; content slightly dense.
PLEUROPSEUDOCYSTIDIA cylindric to slightly tortuous, never emergent, 3.5-4
um wide; content refringent. LAMELLAR edge sterile. CHEILOMACROCYSTIDIA
28-43 x 6.5-9 um, fusiform to lageniform, mostly with moniliform apex.
MARGINAL CELLS 15-35 x 7-10 um, cylindric to clavate, slightly thick-walled,
hyaline. SUBHYMENIUM $20 um thick. HYMENOPHORAL TRAMA with abundant
lactifers. PILEIPELLIS an ixocutis to ixotrichoderm, <320 um thick, composed
of few repent hyaline hyphae and abundant erect hyphae (2-4 um broad).
CLAMP CONNECTIONS absent.

EcoLocy & DISTRIBUTION — Gregarious to caespitose under Abies densa
and Tsuga dumosa in subalpine mixed (coniferous and broadleaf) forest.
August. Fairly common.

Lactarius spp. nov. (India) ... 121

ars.

10 um.

Pate 8. Lactarius pyriodorus (Holotype KD 11-027).

A. Pleuromacrocystidia. B. Marginal cells. Scale bars

122 ... Das, Verbeken, & Nuytinck

PLATE 9. Lactarius pyriodorus (Holotype KD 11-027):.
A. Radial section through pileipellis. Scale bar = 10 um.

Lactarius spp. nov. (India) ... 123

ADDITIONAL SPECIMENS EXAMINED: INDIA. SIKKIM: North District, Dombang,
27°44'08.7”'N 88°45’58.3’E, alt. 2975 m, 14. VIII.2009, A. Verbeken, K. Das & K.V. Putte,
AV-KD-KVP 050 & 051 (BSHC, GENT); alt. 2945 m, 23.VIII.2011, K. Das, KD 11-068
(BSHC); North District, Zema, 27°47’20.0’N 88°32’56.1”E, alt. 3079 m, 31.VIII.2012, K.
Das, KD 12-223, KD 12-226 (BSHC).

Notes — The viscid to glutinous subzonate pileus, ixocutis to ixotrichoderm
nature of the pileipellis, and latex that stains the context and lamellae distinctly
lilac to violet place this species in L. sect. Uvidi. Only one representative of
L. subg. Piperites with lilac milk has been described from Asia (northern
Thailand)—L. formosus H.T. Le & Verbeken (Le et al. 2007). Lactarius formosus
clearly differs from L. pyriodorus because its pileus is completely covered with
bundles of glutinous hairs.

In the ITS-generated phylogenetic tree, the resolution in the group of lilac
staining species is very low, and the Indian species is not distinctly separated
from a cluster with the European species Lactarius luridus (Pers.) Gray and
L. brunneoviolaceus M.P. Christ. We have previously observed that in this
group the morphological variation appears much higher than the ITS
molecular variation. Since we observe clear morphological differences
between L. pyriodorus and the European taxa (also not supported in this tree,
although generally accepted as separate species), we choose to present these
Indian specimens as a new species. Its darker colours and subzonate aspect
are shared with L. luridus, which typically associates with broadleaf trees and
can be separated by its pileus that is spotted but never with thick gluten (as
in L. pyriodorus). Also, in L. luridus, the stipe base is never strigose and the
distinctive sweetish pear-like odour is absent (Heilmann-Clausen et al. 1998).
Lactarius brunneoviolaceus and L. pseudouvidus Kihner are two arctic-alpine
species associated with Salix that differ greatly from our Indian species in their
very small habit. Lactarius brunneoviolaceus is further distinguished by its
cedar-oil odour and rather large spores (8.8-11.9 x 6.7-8.6 um) (Heilmann-
Clausen et al. 1998, Basso 1999). Lactarius pseudouvidus differs microscopically
by its distinctly lower (<0.3 um) spore ornamentation and differently shaped
hymenial cystidia (cylindric to subfusiform with rounded apex) (Heilmann-
Clausen et al. 1998). Lactarius pyriodorus is superficially similar to the European
L. uvidus (Fr.) Fr., which can also occur under coniferous trees, but it never has
such dark colours (Heilmann-Clausen et al. 1998), and its spores have rounded
warts and ridges forming a more incomplete reticulum.

This is for the first time that distinct sweetish odour reminiscent of pears
(also known from the European Inocybe corydalina) has been encountered in
Lactarius.

124 ... Das, Verbeken, & Nuytinck

Lactarius yumthangensis K. Das & Verbeken, sp. nov. PLATE 10-12, 13C,D,F
MycoBank MB 804890

Differs from Lactarius trivialis by its lighter spore print, smaller spores, and pileipellis
ixocutis.

Type: India. Sikkim: North District, Yumthang valley of Shingba Rhododendron
sanctuary, 27°46'51.9”N 88°42’37.6’E, alt. 3586 m, 30.VIIL.2011, K. Das, KD 11-147
(Holotype, BSHC, isotype, GENT).

EryMo_oey: after the name of the locality from where the type specimen was collected

PiLEus 37-98 mm diam., at first convex with inrolled margin, becoming
planoconvex to applanate with depressed indented centre or funnel shaped
at maturity, surface smooth, viscid when dry, slimy when moist, vinaceous
grey (a: 80) or slightly paler (purplish grey), mostly with distinct darker
spots forming zonations; margin non-striate, paler, decurved with maturity.
LAMELLAE broadly adnate to decurrent, close to medium crowded (9-11/cm
at margin), some forked, with lamellulae in 5 series, cream-yellow to yellow
(a: 6F) or ochraceous, sometimes with rusty spots after maturity, unchanging
when bruised; edge entire, concolorous. STIPE 21-50 x 9-22 mm, cylindrical to
ventricose, often constricted below the juncture of the lamellae, surface slightly
longitudinally venose, whitish on constriction, then vinaceous to vinaceous grey
(a: 80), cream yellow towards base, with ochraceous areas/spots towards base
after maturity. CONTEXT yellowish white to cream, hollow, multi-chambered
in stipe, becoming greenish with FeSO, and orange-yellow with KOH. LaTex
chalky white, changing to orange-yellow with KOH, turning pale cream (a: 3C)
after some time, slowly (on drying) greenish yellow (a: 57) after 2 hours (on cut
lamellae). TasTE acrid. ODour fruity. SPORE PRINT pale yellow (c: 30Y, 2M).
BASIDIOSPORES 6.4—7.1-8.0 x 5.3-6.0-6.9 um, (Q = 1.06-1.18-1.32),
subglobose to ellipsoid; ornamentation amyloid, <1 um high, composed of
rather regular, narrow and acute ridges mostly arranged in parallel groups
forming zebroid pattern, short ridges and isolated irregular warts present
between ridges, but, never forming reticulum; plage amyloid distally but
often not distinguishable. Bastp1a 38-58 x 9-11 um, 2-4-spored, clavate to
ventricose; sterigmata long, 5-9 x 2-2.5 um. PLEUROMACROCYSTIDIA 44-90
x 7.5-10 um, abundant, clavate to fusiform (sometimes with mucronate apex),
emergent <36 um; content slightly dense but never needle-like, slightly thick-
walled (wall up to 0.6 um). PLEUROPSEUDOCYSTIDIA irregularly tortuous
with rounded apex, 2.5-4 um wide; contents dense. LAMELLAR EDGE Sterile.
CHEILOMACROCYSTIDIA 27-55 x 8-9 um, subclavate or narrowly clavate to
fusoid, emergent <25 um, slightly thick walled (wall <0.7 um). MARGINAL
CELLS forming chains of elements, terminal cells 11-18 x 5-8 um, subcylindric
to subclavate or clavate, hyaline. SuBHYMENIUM <28 um thick, cellular.

Lactarius spp. nov. (India) ... 125

PLATE 10:. Lactarius yumthangensis (Holotype KD 11-147).
A. Fresh basidiomata showing lamellae and lamellulae. B. Basidiospores.
C. Pleuropseudocystidia. D. Marginal cells. Scale bars: B = 5 um; C, D = 10 um.

126 ... Das, Verbeken, & Nuytinck

Aan
Pea)

2% VV

PLATE 11. Lactarius yumthangensis (Holotype KD 11-147).
A. Pleuromacrocystidia. B. Cheilomacrocystidia. C. Basidia. Scale bars = 10 um.

Lactarius spp. nov. (India) ... 127

PLATE 12. Lactarius yumthangensis (Holotype KD 11-147).
A. Radial section through pileipellis. Scale bar = 10 um.

PILEIPELLIS an ixocutis, composed of narrow repent hyaline hyphae and broad
(<7 um) septate hyphae embedded in a layer of slime. CLAMP CONNECTIONS
absent.

EcoLoGy & DISTRIBUTION — Gregarious to caespitose under Betula utilis
in subalpine mixed (coniferous and broadleaf) forest. August-September.
Uncommon.

ADDITIONAL SPECIMENS EXAMINED: INDIA. Sikkim: North District, Yumthang
valley of Shingba Rhododendron sanctuary, 27°46’51.9”N 88°42’37.6’E, alt. 3586 m,
29.VIII.2011, K. Das, KD 11-150 (BSHC); 27°45’11.1”N 88°43’47.5’E, alt. 3580 m,
01.1X.2011, K. Das, KD 11-172 (BSHC).

Notes —The viscid to slimy zonate pileus and the ixocutis to ixotrichoderm
nature of the pileipellis undoubtedly place the present species in L. subg.

128 ... Das, Verbeken, & Nuytinck

ee 5 I

PLATE 13. Lactarius pyriodorus. A. Fresh basidiomata. B. Latex oozes out from cut lamellae.
E. Basidiospores (SEM). Lactarius yumthangensis. C. Fresh basidiomata. D. Basidiomata showing
constricted stipe-apex. FE Basidiospores (SEM). Scale bars: E = 5 um; F = 3 um.

Piperites. Lactarius yumthangensis can be recognized by its vinaceous grey to
purplish grey pileus, venose stipe with a constricted apex, chalky white latex

Lactarius spp. nov. (India) ... 129

that changes to orange-yellow with KOH and becomes greenish yellow after
long exposure, and growth under Betula sp.

Morphologically, L. flexuosus (Pers.) Gray and L. trivialis (Fr.) Fr. (both
reported from Europe) appear quite similar. However, both can be distinguished
from L. yumthangensis by the slightly darker spore print (c: 40Y, 5M). Moreover,
the white latex of L. flexuosus is unchanging (never becoming greenish yellow
after exposure), the stipe is not constricted at the apex (Kranzlin 2005), and
some cheilomacrocystidia have a moniliform apex (Heilmann-Clausen et al.
1998). In L. trivialis, spores are distinctly larger (7.3-10 x 5.9-7.8 um) and the
pileipellis is an ixocutis to ixotrichoderm (Heilmann-Clausen et al. 1998).

Acknowledgments

The authors are thankful to the Director, Botanical Survey of India, Kolkata (India)
and Department of Forest, Environment and Wild Life Management, Government
of Sikkim, for providing all kinds of facilities during this study. They are grateful to
Prof. N.S. Atri (India) and Dr. Xiang-Hua Wang (China) for reviewing the manuscript
and giving valuable suggestions. Thanks are also due to Ursula Eberhardt, who kindly
provided the sequences for the specimens. Assistance (during macrofungal surveys
to Dombang, Shingba Rhododendron sanctuary, and Zema) rendered by S.K. Rai,
S. Pradhan, R. Giri, R.K. Ram and P. Tamang (BSI, SHRC, Gangtok) is duly acknowledged.

Literature cited

Basso MT. 1999. Lactarius Pers. Fungi Europaei, vol. 7. Mycoflora, Alassio.

Berkeley MJ. 1852. Decades of Fungi: Decades XXXIX., XL. Sikkim and Khassya Fungi. Hooker’s
J. Bot. 4: 130-142.

Buyck B, Hofstetter V, Eberhardt U, Verbeken A, Kauff F. 2008. Walking the thin line between Russula
and Lactarius: the dilemma of Russula sect. Ochricompactae. Fungal Diversity 28: 15-40.

Buyck B, Hofstetter V, Verbeken A, Walleyn R. 2010. Proposal to conserve Lactarius nom. cons.
(Basidiomycota) with a conserved type. Taxon 59(1): 295-296.

Castresana J. 2000. Selection of conserved blocks from multiple alignments for their use in
phylogenetic analysis. Mol. Biol. Evol. 17(4): 540-552.
http://dx.doi.org/10.1093/oxfordjournals.molbev.a026334

Das K. 2009. Mushrooms of Sikkim 1: Barsey Rhododendron Sanctuary. Sikkim State Biodiversity
Board, Gangtok and Botanical Survey of India, Kolkata.

Das K, Verbeken A. 2011. Three new species of Lactarius from Sikkim, India. Cryptogamie
Mycologie 32(4): 365-381. http://dx.doi.org/10.7872/crym.v32.iss4.2011.365

Das K, Verbeken A. 2012. New Species of Lactarius Subg. Plinthogalus and new records of Lactifluus
subg. Gerardii (Russulaceae) from Sikkim, India. Taiwania 57(1): 37-48.

Das K, Van de Putte K, Buyck B. 2010. New or interesting Russula from Sikkim Himalaya (India).
Cryptogamie Mycologie 31(4): 373-387.

Das K, Atri NS, Buyck B. 2013. Three new species of Russula (Russulales) from India. Mycosphere
4(4): 707-717. http://dx.doi.org/10.5943/mycosphere/4/4/9

Gardes M, Bruns TD. 1993. ITS primers with enhanced specificity for Basidiomycetes -
application to the identification of mycorrhizae and rusts. Molecular Ecology 2: 113-118.
http://dx.doi.org/10.1111/j.1365-294X.1993.tb00005.x

130 ... Das, Verbeken, & Nuytinck

Hall TA. 1999. BioEdit: a user-friendly biological sequence alignment editor and analysis program
for Windows 95/98/NT. Nucleic Acids Symposium Series 41: 95-98.

Heilmann-Clausen J, Verbeken A, Vesterholt J. 1998. The genus Lactarius. Fungi of Northern
Europe, vol. 2. The Danish Mycological Society.

Henderson DM, Orton PD, Watling R. 1969. British fungus flora: agarics and boleti: colour
identification chart. Royal Botanic Garden Edinburgh, UK.

Hesler LR, Smith AH. 1979. North American species of Lactarius. USA, Ann Arbor, University of
Michigan Press.

Holmgren PK, Holmgren NH, Barnett LC. 1990. Index Herbariorum. Part 1: Herbaria of the world,
86" ed. Bronx: New York Botanical Garden, USA.

Katoh K, Standley DM. 2013. MAFFT multiple sequence alignment software version 7:
improvements in performance and usability. Molecular Biology and Evolution 30: 772-780.
http://dx.doi.org/10.1093/molbev/mst010

Kornerup A, Wanscher JH. 1981. Methuen handbook of colour. UK, London, EyreMethuen Ltd.
Reprint.

Kranzlin, F. 2005. Fungi of Switzerland. Volume 6, Russulaceae. Verlag Mykologia Luzern,
Switzerland.

Le HT, Nuytinck J, Verbeken A, Desjardin DE, Lumyong S. 2007. Lactarius in Northern Thailand: 1.
Lactarius subg. Piperites. Fungal Diversity 24: 173-224.

Nuytinck J, Verbeken A. 2003. Lactarius sanguifluus versus Lactarius vinosus molecular and
morphological analyses. Mycological Progress 2: 227-234.
http://dx.doi.org/10.1007/s11557-006-0060-5

Stamatakis A. 2006. RAxML-VI-HPC: maximum likelihood-based phylogenetic analyses with
thousands of taxa and mixed models. Bioinformatics 22: 2688-2690.
http://dx.doi.org/10.1093/bioinformatics/btl446

Van de Putte K, Nuytinck J, Stubbe D, Le HT, Verbeken A. 2010. Lactarius volemus sensu lato
(Russulales) from northern Thailand: morphological and phylogenetic species concepts
explored. Fungal Diversity 45: 99-130. http://dx.doi.org/10.1007/s13225-010-0070-0

Van de Putte K, Nuytinck J, Das K, Verbeken A. 2012. Exposing hidden diversity by concordant
genealogies and morphology—a study of the Lactifluus volemus (Russulales) species complex
in Sikkim Himalaya (India). Fungal Diversity 55: 171-194.
http://dx.doi.org/10.1007/s13225-012-0162-0

Verbeken A, Horak E. 2000. Lactarius (Basidiomycota) in Papua New Guinea. 2. Species in tropical-
montane rainforest. Austr. Syst. Bot. 13: 649-707. http://dx.doi.org/10.1071/SB99023

Verbeken A, Nuytinck J. 2013. Not every milkcap is a Lactarius. Scripta Botanica Belgica 51:
162-168.

White TJ, Bruns T, Lee SS, Taylor J. 1990. Amplification and direct sequencing of fungal ribosomal
RNA genes for phylogenetics. 315-322, in: MA Innis et al. (eds). PCR Protocols: a guide to
methods and applications. Academic Press, New York.

MYCOTAXON

http://dx.doi.org/10.5248/130.131
Volume 130, pp. 131-143 January-March 2015

Synonymy of two species of Bipolaris
from aquatic crops of Poaceae

ZI-LAN XIAO’, KEVIN D. Hype’, & JING-ZE ZHANG"

"Institute of Biotechnology, College of Agriculture & Biotechnology, Zhejiang University,
Hangzhou 310058 China

?Institute of Excellence in Fungal Research, School of Science, Mae Fah Luang University,
Chiang Rai 57100 Thailand

*CORRESPONDENCE TO: jzzhang@zju.edu.cn

ABSTRACT— Our morphological data indicate that conidia of Bipolaris zizaniae (a pathogen
causing brown spot on leaves of Zizania latifolia) are similar to those of B. oryzae. Our
sequences of the rDNA ITS, GPDH, and EF1-a gene regions from B. zizaniae and blast
searches revealed a 99-100% similarity with sequences of B. oryzae. Phylogenetic studies
also cluster B. zizaniae isolates with B. oryzae in a clade with 100% bootstrap support.
Pathogenicity testing also confirmed that B. zizaniae does infect both Zizania latifolia and
rice, causing brown spots.

Key worps—graminicolous fungi, molecular phylogeny, taxonomy

Introduction

Zizania latifolia (Griseb). Turcz. ex Stapf is a perennial grass parasitized by
the smut fungus Ustilago esculenta Henn., which causes swelling of its upper
culms. The swollen culms are used as an aquatic vegetable, commonly called
jiaobai (white bamboo) in China (Zhang et al. 2012). Due to its unique flavor
and delicacy, the demand for jiaobai has steadily increased in tandem with
the rapid economic development of China. Zhejiang Province has become
the largest Z. latifolia cultivation area in China (Xu et al. 2009). Brown spot
is one of the most severe leaf spot diseases of this crop plant. With large-scale
cultivation and changed growing conditions, brown spot incidence has become
more severe in the fields, and the disease has become a severe problem in
crop production. The pathogen causes a large number and areas of coalescing
spots (especially during seedling production in greenhouses), and these spots
expand or coalesce to form large lesions, eventually resulting withering and
death of entire leaves (Fic. 1A-D).

132 ... Xiao, Hyde, & Zhang

The pathogen causing this disease was first described as Helminthosporium
zizaniae by Nisikado (1929). Although H. zizaniae resembles H. oryzae
morphologically and by infecting Oryza sativa L. [rice], Nisikado (1929)
differentiated the two species based on the shape of the conidial base, conidial
dimensions, and conidial hilum. Shoemaker (1959) transferred H. zizaniae and
H. oryzae to Bipolaris, a placement later accepted by Shoemaker (1966) and
Sivanesan (1987). Sivanesan (1987) used conidial dimensions as an important
character in his dichotomous key to Bipolaris; he distinguished B. zizaniae
as having >150 um long conidia compared with the <150 um long conidia
in B. oryzae. However, conidial dimensions are usually variable in Bipolaris
(Sivanesan et al. 1987; Manamgoda et al. 2012), and the conidial length in
H. oryzae is not stable and can be more than 150 um in some strains (Chang
1978). Other studies showed that B. oryzae is interfertile with B. zizaniae and
that their isolates from rice and Zizania are reciprocally infective (Chang 1974;
Tsuda & Ueyama 1975, 1976). Bipolaris zizaniae and B. oryzae are now regarded
as conspecific (Manamgoda et al. 2014).

Accurate identification and precise naming of species are crucial, for species
names provide an ideal framework for storage and information retrieval for
effective disease management (Inderbitzin et al. 2011, Manamgoda et al. 2012).
Bipolaris zizaniae is an important pathogen on Z. latifolia, but due to limited
distribution, there is little information on its effects; even the morphological
descriptions of the pathogen are inadequate (Nisikado 1929, Sivanesan 1987). In
contrast, B. oryzae [sexual state previously known as Cochliobolus miyabeanus]
causes brown spots on leaves and glumes of rice, leading to serious losses in
yield (Scheffer 1997). Thus this pathogen and rice disease have been studied
worldwide (Kulkarni et al. 1980, Kubo et al. 1989, Amadioha 2002, Manandhar
et al. 1998, Xio et al. 1991). Nonetheless, the relationship between B. zizaniae
and B. oryzae is unresolved. With the use of molecular taxonomy, DNA
sequence analyses allow testing the morphological species circumscriptions
and providing new insights into species relationships, especially for complexes
of closely related, morphologically similar species (Manamgoda et al. 2012,
2014; Udayanga et al. 2012; Hyde et al. 2014). Studies of Bipolaris species have
already revealed relationships between morphological species circumscriptions
and molecular data, but there has been no study of B. zizaniae (Manamgoda et
al. 2011, 2012, 2014; Hyde et al. 2014).

We used sequences from the ITS (internal transcribed spacer), GPDH
(glyceraldehyde-3-phosphate dehydrogenase), and EFl-a (translation
elongation factor 1-a) gene regions to investigate the phylogenetic relationships
of six Bipolaris cultures isolated from Zizania latifolia in southeast China.
The objectives were to review morphological similarities in B. zizaniae and
B. oryzae and to resolve their relationship.

Bipolaris zizaniae confirmed as a synonym of B. oryzae ... 133

FiGurE 1 Symptoms of Bipolaris brown spot on Zizania latifolia and Oryza sativa. A-D. On
nature host of Z. latifolia. A. Early spots. B. Severe spots. C. Removed leaves with severe brown
spots from seedlings in the greenhouse. D. Leaf blight. E-E Symptom variability on backs of
leaves of Z. latifolia after artificial inoculation. E. Five days after inoculation. F Eighteen days
after inoculation. G-I. Symptom variability on leaves of O. sativa after artificial inoculation.
G. Brown spots eight days after inoculation. H. Brown spots on rear of leaf nine days after
inoculation. I. Brown spots twelve days after inoculation.

134 ... Xiao, Hyde, & Zhang

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Bipolaris zizaniae confirmed as a synonym of B. oryzae ... 135

Materials & methods

Collection and isolation of fungi

Diseased leaves of Zizania latifolia with brown spots growing in fields or greenhouses
were collected and returned to the laboratory. Tissue pieces (each approximately 5 x 5
mm) were cut from the margins of brown spots, between healthy and infected parts of
leaves, which were previously surface disinfected with 0.5% NaOCl. The pieces were
plated on to PDA medium and incubated at 25°C for 7 days. Mycelia from the infected
samples were transferred to fresh PDA medium. Single-spore isolation was performed
as described by Chomnuntiet al. (2014). Agar discs (with mycelium on the surface, each
approximately 5 x 5 mm) from these isolates were stored in 1.5 mL cryotubes with 20%
glycerol at -70°C.

Herbarium materials are deposited at MFLU herbarium of Mae Fah Luang University,
Chiang Rai, Thailand (MFLU). The cultures are maintained at Mae Fah Luang University
Culture Collection (MFLUCC) and Institute of Biotechnology, Zhejiang University,
Zhejiang Province, China (ZJU).

DNA extraction

Genomic DNA was extracted using a modified protocol of Zhang and Li (2009).
Fungal isolates were cultured on PDA at 25°C for 7 days. Fresh mycelia were scraped
from the surfaces of PDA plate and grounded to a fine powder in liquid nitrogen. 500 mg
power was transferred to a 1.5 mL microcentrifuge tube and 700 uL of preheated (60°C)
2x CTAB extraction buffer (2% hexadecyltri-methlammonium bromide(w/v), 100 mM
Tris-HCl, 1.4 M NaCl, 30 mM EDTA, pH 8.0) was added. The solution was incubated
in a water bath at 65°C for 60 min with a gentle swirling. Following a phenol/chloroform
extraction, the genomic DNA was precipitated by isopropanol in the presence of sodium
acetate. The precipitate was rinsed with 70% ethanol centrifuged at 12,000 rpm and
genomic DNA was resuspended in 50 uL TE buffer and stored in —20°C.

PCR amplification

The DNA fragments were amplified in an automated thermal cycler (Eppendorf AG,
Germany). Amplification was performed in a 50 uL reaction volume which contained
5 wL 10x PCR buffer, 1 uL of each primer (10 uM), 2 uL template DNA, and 0.5 uL Taq
DNA polymerase. Primers ITS6 and ITS4 (Zhang & Li. 2009) were used to amplify
the ITS and 5.8 S regions. The thermal cycling program was performed with 35 cycles
after an initial denaturation at 95°C for 4 min. Each cycle included a denaturation step
at 95°C for lmin, an annealing step at 55°C for lmin, and an extension step at 72°C
for 1.5 min. The primers gpd1/ gpd2 (Berbee et al. 1999) were used for amplifying the
GPDH gene with the changes of annealing temperature 52°C. Similarly, the EF-la gene
was amplified using primers EF 983/2218R (Schoch et al. 2009) with the changes of
annealing temperature 54°C.

Sequence alignment and phylogenetic analysis

The purified PCR products were submitted to Sunny Biotechnology Company
Limited (Shanghai, China) for sequencing in both directions. Sequence files were
assembled and edited using BioEdit software (Hall 1999). A blast search was performed

136 ... Xiao, Hyde, & Zhang

for searching the highly similar sequences in the GenBank database and related
sequences were downloaded (TABLE 1). The sequences were aligned with Clustal X 25
(Thompson et al. 1997). Sequences were analyzed phylogenetically in PAUP v4.0b10
(Swofford 2002) and trees were visualized with TreeView (Page 1996). Phylogenetic
trees were inferred from the ITS, GDPH and EF1-a sequence data set using parsimony
analysis with all characters weighted equally and all gaps treated as missing data.

Morphological studies

Conidia and conidiophores taken directly from nature hosts were examined in 3%
KOH, using a Zeiss Axiophot 2 microscopy with Axiocam CCD camera and AxioVision
digital imaging software (AxioVision Software Release 3.1.v.3-2002; Carl Zeiss Vision
Imaging Systems). Cultural characteristics and morphology were determined on PDA,
and water agar and wheat straw (WSA) [2 % water agar (10 g/L) with autoclaved wheat
straw placed onto the medium] and incubated for 7-10 days at 25°C under 12/12 h
alternation of near-UV light. 20 uL conidial suspension (10* conidia/mL) were dropped
on the WA (water agar) medium at 25°C for 24 h and characteristics of conidia
germination were observed.

Scanning electron microscopy

The 0.5 x 0.5 mm’ pieces from diseased spots with conidia and conidiophores were
placed in glutaraldehyde (2.5% vol/vol) for 48 h at 4°C. Samples were washed three times
in 0.1 M sodium phosphate buffer (pH 7.0) and post-fixed in osmium tetroxide (1% wt/
vol) for 1.5 h at 20°C. Samples were rinsed thoroughly with 0.2 M phosphate buffer (pH
6.8) and dehydrated in a graded ethanol series (30, 50, 70, 80, 90, 95, and 100%). Pieces
were embedded in Spurr’s epoxy resin. The sites containing conidia were identified
by light microscopy and ultrathin sections were cut with a Reichert-Jung Ultracut E
Ultramicrotome with a diamond knife. Ultrathin sections were collected on Formvar-
coated slot copper grids; after drying, the grids were stained with uranyl acetate and
lead citrate and examined using a transmission electron microscope (Hitachi S-3000N,
Tokyo, Japan).

Pathogenicity Tests

Healthy field cuttings of Z. latifolia with 3-5 nodes were cultivated in a 25-cm diam.
barrel with cultivated soil and placed in greenhouse at 26-28°C for one month. Oryza
sativa seedlings grown in seedbeds were transplanted into a 20-cm diam. barrel and
placed in greenhouse under the same conditions for one month.

An aqueous inoculation suspension (1 x 10° conidia/mL) with 1% vol/vol Tween-20
was prepared from 8-10-day old cultures. Pathogenicity tests were conducted on healthy
leaves of Zizania latifolia and Oryza sativa. A 50 uL conidial suspension was dropped on
two sides of each leaf and then inoculated plants were covered by large plastic bags and
incubated at 27-28°C for 48 h after which the plastic bags were removed. Control leaves
were inoculated with 50 uL sterile water. Symptoms and sizes of diseased spots were
observed and recorded each day. The fungus was re-isolated by cutting small portions
from the margin of diseased spots; these were surface sterilized and placed on PDA
plates. The experiments were repeated twice to confirm the results.

Bipolaris zizaniae confirmed as a synonym of B. oryzae ... 137

Results

Phylogenetic analysis

Isolates of Bipolaris were obtained from Z. latifolia and purified, and six
single-spore isolates were selected for study. Six strains of Bipolaris from
Z. latifolia had identical ITS and GPDH gene sequences, but in the EF-la
gene sequence there was one base difference between the two strains from
Hangzhou (MFLUCC13-0511, MFLUCC13-0512) and the other four strains
(MFLUCC13-0513, MEFLUCC13-0514, MFLUCC13-0515, MFLUCC13-0516). A
GenBank search showed that there were no sequence data submitted for B.
zizaniae, but a blast search of our ITS, GPDH, and EFl-a sequences from

Bipolaris nodulosus CBS 160.58
65 Bipolaris peregianensis BRIP 12790
400 99 | Bipolaris microlaenae CBS 280.91
Bipolaris microlaenae BRIP 15613
Bipolaris cynodontis |CMP 6128
Bipolaris oryzae MFLUCC 10-0715
Bipolaris oryzae MFLUCC 10-0694 | ftom rice
100| Bipolaris oryzae MFLUCC 10-0733
100 Bipolaris oryzae MFLUCC 13-0511
Bipolaris oryzae MFLUCC 13-0512
Bipolaris oryzae MFLUCC 13-0513 |om zizania
61 Bipolaris oryzae MFLUCC 13-0516
/7' | Bipolaris oryzae MFLUCC 13-0515
67 | Bipolaris oryzae MFLUCC 13-0514
Curvularia graminicola BRIP 23186a
86 Curvularia homomorpha CBS 156.60
Curvularia coicis CBS 192.29
Curvularia tripogonis BRIP 12375
92 Curvularia ovariicola BRIP 15882
ius Curvularia ravenelii BRIP 13165
Curvularia heteropogonis CBS 284.91
400 pCurvularia lunata MFLUCC 10-0706
Curvularia lunata CBS 730.96
60 Curvularia australiensis CBS 172.57
“ Curvularia ellisii CBS 193.62
Curvularia Spicifer CBS 274.52
400 r- Curvularia hawaiiensis BRIP 10971
Curvularia hawaliensis CBS 173.57
Curvularia tuberculata CBS 146.63

_10

FiGuRE 2. Phylogram generated from parsimony analysis based on ITS, GPDH and EF 1-a
sequence data of Bipolaris and Curvularia species. Data were analysed with random sequence
addition, unweighted parsimony and by treating gaps as missing data. Bootstrap values =>50% are
indicated. Species names are followed by the number of the deposited voucher. The tree is rooted
with Curvularia tuberculata.

138 ... Xiao, Hyde, & Zhang

B. zizaniae showed a 99-100% similarity with those of B. oryzae lodged in
GenBank. Of the 63 reliable or voucher B. oryzae ITS sequences in GenBank,
11 were identical with B. zizaniae and 41 differed by 1-2 bases. Similarly, of
the 16 reliable or voucher B. oryzae GPDH sequences in GenBank, one was
identical with B. zizaniae and 15 differed by 1-2 bases. Finally, of five B. oryzae
EF1-a sequences (Manamgoda et al. 2012), four were identical with B. zizaniae
and one differed by 1-2 bases.

A parsimony tree was constructed from the ITS, GPDH, and EF1-a data
from 29 fungal isolates (TABLE 1). The four Bipolaris isolates from Z. latifolia
with identical sequences grouped together with another two Bipolaris isolates
from Z. latifolia (with one base difference) with 61% bootstrap value support
(Fic. 2). This group clustered with three B. oryzae isolates with a 100% bootstrap
support. Fic. 2 clearly shows that the Bipolaris isolates from Z. latifolia are
inseparable from B. oryzae. Sequence variation between Bipolaris isolates
from Z. latifolia and B. oryzae isolates was less than interspecific variation.
Six Bipolaris isolates from Z. latifolia and three B. oryzae isolates clustered
with B. coffeana Sivan. to form a clade with a 70% bootstrap value support
(Fic. 2). However, the B. coffeana subclade (based on a single culture) and the
B. oryzae/B. zizaniae subclade were separated by >40 base differences, indicating
that they are not conspecific.

The description below is based on our collections of Bipolaris isolates from
Z. latifolia. Except for size variation, these characters are almost identical to
those from the type specimen from the same host from Japan (Nisikado 1929).
The pathogen causes brown spots on leaves of Zizania latifolia and infects
almost all aerial parts of the plant.

Bipolaris oryzae (Breda de Haan) Shoemaker, Can. J. Bot. 37: 883 (1959) Fig23

= Helminthosporium oryzae Breda de Haan, Bulletin Inst. Bot. Buitenzorg 6: 11 (1900)

= Ophiobolus miyabeanus S. Ito & Kurib., Ann. Phytopath. Soc. Japan 2(1):

7 (1927)
= Cochliobolus miyabeanus (S. Ito & Kurib.) Drechsler ex

Dastur, Indian. J. Agr.Res. 12: 733 (1942)

= Helminthosporium zizaniae Y. Nisik., Ber. Ohara Inst.
Landw. Forsch. Kurashiki 4: 122 (1929)
= Bipolaris zizaniae (Y. Nisik.) Shoemaker, Can. J. Bot. 37: 885 (1959)

Leaf spots dark brown with a paler centre 0.3-0.5 x 0.3-0.6 cm. On a natural
host, conidiophores single or occasionally fasciculate in small groups, rarely
branched, geniculate or straight in sterile part, then becoming slightly to
distinctly geniculate, cicatrized with scars often inflated at base, medium
olivaceous-brown below, paler at the apex, multiseptate, 106-395 um long,
swollen to 7-9 um diam at base, then narrowing to 6-7 um diam (middle)

Bipolaris zizaniae confirmed as a synonym of B. oryzae ... 139

FiGurE 3. Bipolaris oryzae (isolates from Z. latifolia). A-D. Light microscope micrographs.
A. Conidia. B. Conidia on WSA. C. Conidiophore. D. Germinated conidium with one germ tube at
each end. Basal germ tube direction of growth (left below) is semiaxial, close to hilum. E-G. SEM
micrographs. E. Conidia. F. Hilum. G. Germinated conidium with two polar germ tubes. Scale bars:
A, B, D(lower) = 10 um; C, D(upper) = 20 um; E= 15 um; F=5 um; G= 25 um.

and 3-5 um (apex). Conidia (65-)78-110(-131) x (15-)17-28(-34) um,
6-10 distoseptate, olivaceous brown or yellowish brown, obclavate, navicular,
straight or slightly curved, tapering towards the apex, hilum dark, conspicuous,
sometimes slightly protruding. Conidia germinate by the production of two
polar germ tubes. The sexual morph is not found in nature.

On WSA, conidiophores 212-646 x 5.5-10.5 um, olivaceous-brown, paler
at the apex. Conidia brown or dark brown, obclavate, fusoid, often curved,
(73-)80-125(-130) x (1-)18-28(-31) um, 6-11-distoseptate, hilum dark,
conspicuously protruding or truncate.

MATERIALS EXAMINED: CHINA, ZHEJIANG PROVINCE, Hangzhou Suburbs,
September 2012, Zilan Xiao (MFLU13-0092; cultures MFLUCC13-0511 = ZJU0101,
MFLUCC13-0512 = ZJU0102); ANHUI PROVINCE, Yuexi County, September 2012,
Zilan Xiao (MFLU13-0093; cultures MFLUCC13-0513 = ZJU0201, MFLUCC13-0514

= ZJU0202); Fus1AN PROVINCE, Putian city, October 2012, Zilan Xiao (MFLU13-0094;
cultures MFLUCC13-0515 = ZJU0301, MFLUCC13-0516 = ZJU0302).

140 ... Xiao, Hyde, & Zhang

TABLE 2. Synopsis of conidial size and number of distosepta in Bipolaris oryzae from
Zizania latifolia from different origins.

ISOLATE LENGTH (um) WIDTH (um) DISTOSEPTA
ON HosTS MEFLU 13-0092 99 +11.5 25 + 3.0 (5-)7-9
MEFLU 13-0093 87 + 8.7 20 + 3.6 (5-)7-10
MEFLU 13-0094 92 + 10.6 2343.5 (5-)6-10
On WSA MFLUCC 13-0511 96 +11 24+3.8 6-9(-11)
MFLUCC 13-0513 86 + 7.8 19+5.4 6-10
MFLUCC 13-0515 90 + 5.6 2146.7 6-10(-11)

CoMMENTS—Classification in the genus Bipolaris was previously based
on morphology, with conidial characters being considered as important in
species delimitation. In this study conidia of Bipolaris isolates from Z. latifolia
are shorter (78-110 um) and wider (17-28 um) than those reported in the
protologue (25-166 x 12-13 um; Nisikado 1929). Although there is some
variation in conidial size and distosepta in Bipolaris isolates from Z. latifolia
from different locations in China (TaBLE 2), the morphological differences are
not significant. We found it impossible to distinguish B. oryzae from B. zizaniae
based on morphological characters.

Morphology and molecular phylogeny confirm that Bipolaris zizaniae and
B. oryzae are conspecific. Bipolaris oryzae is the correct name for this expanded
taxon because the epithet oryzae (published in 1900) has priority over zizaniae
(published in 1929). We follow Manamgoda et al. (2012, 2014) in preferring
Bipolaris to the earlier generic name Cochliobolus (proposed for the sexual
state).

Pathogenicity testing

The isolates of Bipolaris isolated from Z. latifolia infected the leaves of
Z. latifolia. At 48 h after inoculation, Bipolaris isolates from Z. latifolia induced
small flecks or expanded spots on the lower surface of Z. latifolia leaves.
Symptoms were similar to those observed in the wild (Fic. 1£,F), and no
disease was found on upper surface of inoculated or control leaves. Five days
after inoculation, diseased spots increased in size (Fic. 1E). Symptoms of leaf
blight occurred 18 days after inoculation (Fic. 1F). During the same period,
disease symptoms did not develop on the upper surface or on control leaves.

Similarly, isolates of Bipolaris isolated from Z. latifolia also infected rice
leaves. Small flecks occurred on the upper surface of inoculated leaves after
five days and similar flecks occurred on the lower surface of leaves after six
days. After 8-9 days, diseased spots increased more rapidly in size on the upper
surface of leaves than on the lower surface of leaves (Fic. 1G, H). The diseased
spots were approximately 0.5 cm in diameter 12 days after inoculation (Fie. 11).

Bipolaris zizaniae confirmed as a synonym of B. oryzae... 141

Discussion

Zizania latifolia, which originated in the northern region of China, was
once used as an important grain in ancient times. Cultivated Z. latifolia was
disseminated to Taihu Lake from north to south, and the Taihu Lake basin
is the region associated with the origin of jiaobai (Xu et al. 2008). Jiaobai
cultivation was introduced to eastern and southeastern Asia, including the
Russian Far East, Japan, and Korea (Thrower & Chan 1980, Guo et al. 2007).
It is possible that brown spot was also introduced to the jiaobai-growing areas
with the planting material.

Zizania latifolia commonly grows in shallow water and can be found at the
edges of lakes, ponds, wetlands, and flooded fields (Thrower & Chan 980, Guo
et al. 2007). In some cases, Z. latifolia is able to grow in deeper water (Yamasaki
& Tangel981), because it has a well-developed ventilation system (Yamasaki
1984, 1987). Thus, Z. latifolia growing conditions are similar to that of rice.
In the jiaobai-growing areas, Z. latifolia is usually grown in paddy fields or
in rotation with rice. The fact that pathogenic brown spot of Z. latifolia also
infects rice may be a result of long-term evolution. In morphological taxonomy,
phenotypic plasticity and homoplasy may influence fungal species delimitation
(Pino-Bodas et al. 2011). Bipolaris zizaniae and B. oryzae have previously been
delineated based on morphology, although species boundaries are not clear
(Chang 1974; Tsuda & Ueyama 1975, 1976). Bipolaris zizaniae can be crossed
with B. oryzae to form the sexual state, and both species can infect Zizania
(Chang 1974, Tsuda et al. 1975). The data presented here and in previous studies
suggest that B. oryzae and B. zizaniae are synonymous. Our gene sequence data
analysis shows unequivocally that B. oryzae and B. zizaniae are conspecific.

Acknowledgments

The authors are grateful to Dimuthu $. Manamgoda (School of Science, Mae Fah
Luang University, Thailand) and Lei Cai (State Key Laboratory of Mycology, Institute
of Microbiology, Chinese Academy of Sciences, People’s Republic of China) for their
helpful comments on this manuscript. The research was funded by the National Natural
Science Foundation of China (No. 31070123) and the Special Fund for Agro-scientific
Research in the Public Interest of China (No. 201003004). KD Hyde thanks MFLU grant
No. 56101020032 for funds to study Dothideomycetes.

Literature cited

Amadioha AC. 2002. Fungi toxic effects of extracts of Azadirachta indica against Cochliobolus
miyabeanus causing brown spot disease of rice. Archives of Phytopathology and Plant
Protection 35: 37-42. http://dx.doi.org/10.1080/032354002 1000009597

Berbee M, Pirseyedi M, Hubbard S. 1999. Cochliobolus phylogenetics and the origin of known,
highly virulent pathogens, inferred from ITS and glyceraldehyde-3-phosphate dehydrogenase
gene sequences. Mycologia 91: 964-977. http://dx.doi.org/10.2307/3761627

142 ... Xiao, Hyde, & Zhang

Chang HS. 1974. Intercross fertility between Helminthosporium oryzae, H. zizaniae and an
unidentified Helminthosporium sp. on Zizania aquatica. Botanical Bulletin of Academia Sinica
15: 103-111.

Chang HS. 1978. A elongated conidium strain and mating type distribution of Cochliobolus
miyabeanus. Botanical Bulletin of Academia Sinica 19: 139-144.

Chomnunti P, Hongsanan S, Aguirre-Hudson B, Tian Q, PerSoh D, Dhami MK, Alis AS,
Xu J, Liu X, Stadler M, Hyde KD. 2014. The sooty moulds. Fungal Divers 66:1-36
http://dx.doi.org/10.1007/s13225-014-0278-5

Guo HB, Li SM, Peng J, Ke WD. 2007. Zizania latifolia Turcz. cultivated in China. Genetic Resources
and Crop Evolution 54: 1211-1217. http://dx.doi.org/10.1007/s10722-006-9102-8

Hall TA. 1999. Bioedit: a user-friendly biological sequence alignment editor and analysis program
for Windows 95/98/NT. Nucleic acids symposium series 41: 95-98.

Hyde KD, Nilsson RH, Alias SA, Ariyawansa HA, Blair JE, Cai L, de Cock AWAM, Dissanayake
AJ, Glockling SL, Goonasekara ID, Gorczak M, Hahn M, Jayawardena RS, van Kan JAL,
Laurence MH, Lévesque CA, Li X, Liu JK, Maharachchikumbura SSN, Manamgoda DS,
Martin FN, McKenzie EHC, McTaggart AR, Mortimer PE, Nair PVR, Pawlowska J, Rintoul
TL, Shivas RG, Spies CFJ, Summerell BA, Taylor PWJ, Terhem RB, Udayanga D, Vaghefi N,
Walther G, Wilk M, Wrzosek M, Xu JC, Yan JY, Zhou N. 2014. One stop shop: backbones
trees for important phytopathogenic genera: I (2014). Fungal Divers. 67(1): 21-125.
http://dx.doi.org/10.1007/s13225-014-0298-1

Inderbitzin P, Bostock RM, Davis RM, Usami T, Platt HW, Subbarao KV. 2011. Phylogenetics and
taxonomy of the fungal vascular wilt pathogen Verticillium, with the descriptions of five new
species. PLoS ONE 6: e28341. http://dx.doi.org/10.1371/journal.pone.0028341

Kubo Y, Tsuda M, Furusawa I, Shishiyama J. 1989. Genetic analysis of genes involved in
melanin biosynthesis of Cochliobolus miyabeanus. Experimental mycology 13: 77-84.
http://dx.doi.org/10.1016/0147-5975(89)90010-8

Kulkarni S$, Ramakrishnan K, Hegde RK. 1980. Ecology, epidemiology, and supervised control of
rice brown leaf. International Rice Research Newsletter 5: 13-14.

Manamgoda DS, Cai L, Bahkali AH, Chukeatirote E, Hyde KD. 2011. Cochliobolus: an overview
and current status of species. Fungal Diversity 51: 3-42.
http://dx.doi.org/10.1007/s13225-011-0139-4

Manamgoda DS, Cai L, McKenzie EHC, Crous PW, Madrid H, Chukeatirote E, Shivas RG, Tan YP,
Hyde KD. 2012.A phylogenetic and taxonomic re-evaluation of the Bipolaris - Cochliobolus -
Curvularia complex. Fungal Diversity 56: 131-144.
http://dx.doi.org/10.1007/s13225-012-0189-2

Manamgoda DS, Rossman AY, Castlebury LA, Crous PW, Madrid H, Chukeatirote E, Hyde KD.
2014. The genus Bipolaris. Studies in Mycology 79: 221-288.
http://dx.doi.org/10.1016/j.simyco.2014.10.002

Manandhar HK, Jorgensen HJL, Mathur SB, Smedegaard-Petersen V. 1998. Suppression of rice
blast by preinoculation with a virulent Pyricularia oryzae and the non-rice pathogen Bipolaris
sorokiniana. Phytopathology 88: 735-739. http://dx.doi.org/10.1094/PHY TO.1998.88.7.735

Nisikado Y. 1929. Studies on the Helminthosporium diseases of Gramineae in Japan. Berichte des
Ohara Instituts fir Landwirthschaftliche Forschungen in Kurashiki 4: 111-126.

Page RDM. 1996. Treeview. An application to display phylogenetic trees on personal computer.
Computer Applications in the Biological Sciences 12: 357-358.

Pino-Bodas R, Rosa Burgaz A, Martin MP, Lumbsch HT. 2011. Phenotypical plasticity and homoplasy
complicate species delimitation in the Cladonia gracilis group (Cladoniaceae, Ascomycota).
Organisms Diversity & Evolution 11: 343-355. http://dx.doi.org/10.1007/s13127-011-0062-2

Bipolaris zizaniae confirmed as a synonym of B. oryzae... 143

Scheffer RP .1997. The nature of disease in plants. Cambridge University Press.

Schoch C, Crous PW, Groenewald J, et al. 2009. A class-wide phylogenetic assessment of
Dothideomycetes. Studies in Mycology 64: 1-15. http://dx.doi.org/10.3114/sim.2009.64.01

Shoemaker RA. 1959. Nomenclature of Drechslera and Bipolaris, grass parasites segregated from
Helminthosporium. Canadian Journal of Botany 37: 879-887. http://dx.doi.org/10.1139/b59-073

Shoemaker RA. 1966. A pleomorphic parasite of cereal seeds, Pyrenophora semeniperda. Canadian
Journal of Botany 44: 1451-1456. http://dx.doi.org/10.1139/b66-160

Sivanesan A. 1987. Graminicolous species of Bipolaris, Curvularia, Drechslera, Exserohilum and
their teleomorphs. CAB International.

Swofford D. 2002. PAUP 4.0 b10: Phylogenetic analysis using parsimony. Sinauer Associates,
Sunderland.

Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG. 1997 The CLUSTAL_X windows
interface: flexible strategies for multiple sequence alignment aided by quality analysis tools.
Nucleic Acids Research 25: 4876-4882. http://dx.doi.org/10.1093/nar/25.24.4876

Thrower LB, Chan YS. 1980. Gau sun: a cultivated host-parasite combination from China.
Economic Botany 34: 20-26. http://dx.doi.org/10.1007/BF02859552

Tsuda M, Ueyama A. 1975. Identity of Helminthosporium-leaf spot fungi attacking Oryzoideae
plants growing in Japan Islands (preliminary note). Transaction of the Mycological Society of
Japan 16: 93-94.

Tsuda M, Ueyama A. 1976. Distribution of two mating types of Cochliobolus miyabeanus in field
and a laboratory attempt to produce hybrids between isolates of Helminthosporium oryzae from
the U.S.A. and Japan. Annals of the Phytopathological Society of Japan 42: 7-11.
http://dx.doi.org/10.3186/jjphytopath.42.7

Udayanga D, Liu X, Crous PW, McKenzie EH, Chukeatirote E, Hyde KD. 2012. A multi-
locus phylogenetic evaluation of Diaporthe (Phomopsis). Fungal Diversity 56: 157-171.
http://dx.doi.org/10.1007/s13225-012-0190-9

Xio JZ, Tsuda M, Doke N, Nishimura S. 1991. Phytotoxins produced by germination spores of
Bipolaris oryzae. Phytopathology 81: 58-64. http://dx.doi.org/10.1094/Phyto-81-58

Xu C, Hu MH, Guo DP. 2009. Current status and prospects of aquatic vegetable industry in
Zhejiang province. ChangJiang Veget 16: 106-109.

Xu X, Ke W, Yu X, Wen J, Ge S. 2008. A preliminary study on population genetic structure
and phylogeography of the wild and cultivated Zizania latifolia (Poaceae) based on ADH1A
sequences. Theoretical and Applied Genetics 116: 835-843.
http://dx.doi.org/10.1007/s00122-008-0717-3

Yamasaki S. 1984. Role of plant aeration in zonation of Zizania latifolia and Phragmites australis.
Aquatic Botany 18: 287-297. http://dx.doi.org/10.1016/0304-3770(84)90070-6

Yamasaki S. 1987. Oxygen demand and supply in Zizania latifolia and Phragmites australis. Aquatic
Botany 29: 205-215. http://dx.doi.org/10.1016/0304-3770(87)90016-7

Yamasaki S, Tange I. 1981. Growth responses of Zizania latifolia, Phragmites australis
and Miscanthus sacchariflorus to varying inundation. Aquatic Botany 10: 229-239.
http://dx.doi.org/10.1016/0304-3770(81)90025-5

Zhang JZ, Li MJ. 2009. A new species of Bipolaris from the halophyte Sesuvium portulacastrum
in Guangdong Province, China. Mycotaxon 109: 289-300. http://dx.doi.org/10.5248/109.289

Zhang JZ, Chu FQ, Guo DP, Hyde KD, Xie GL. 2012. Cytology and ultrastructure of interactions
between Ustilago esculenta and Zizania latifolia. Mycological Progress 11: 499-508.
http://dx.doi.org/10.1007/s11557-011-0765-y

MY COTAXON

http://dx.doi.org/10.5248/130.145
Volume 130, pp. 145-164 January-March 2015

Myxomycetes of Sonora (Mexico) 6.
Central plains of the Sonoran Desert

Marcos LIZARRAGA ', GABRIEL MORENO ?,, MARTIN ESQUEDA 3,
CYNTHIA SALAZAR-MARQUEZ 3, & MARTHA L. CORONADO 4

' Dpto. Ciencias Quimico Biologicas, Instituto de Ciencias Biomédicas, Univ. Autonoma de Ciudad Judrez,
Anillo Envolvente Pronaf y Estocolmo s/n, Ciudad Juarez, Chihuahua 32300, México

? Dpto. Ciencias de la Vida, Edificio de Biologia, Univ. Alcala, Madrid, 28805, Espana

> Centro de Investigacion en Alimentacion y Desarrollo,
A.C. Apartado Postal 1735, Hermosillo, Sonora, 83304, México

‘Univ. Estatal de Sonora. Apartado Postal 11, Admon. 11, Hermosillo 83000, Sonora, México

* CORRESPONDENCE TO: gabriel.moreno@uah.es

ABSTRACT — Myxomycetes from 10 localities in the central plains of the Sonoran desert were
studied using the moist chamber culture technique. Thirty-three species have been identified,
of which five (Comatricha mirabilis, Cribraria confusa, Didymium eremophilum, Perichaena
luteola, and P. stipitata) represent new records for Sonora, and Comatricha mirabilis is cited
for the first time from Mexico. Light and scanning electron microphotographs illustrate the
most diagnostic characters of these species, which are rare or scarcely mentioned in the
literature.

Key worps — Amoebozoa, chorology, myxobiota, slime moulds, taxonomy

Introduction

The Sonoran desert, which lies in northwestern Mexico and the southwestern
United States, covers a 260,000-km2 area, with the largest portion occurring in
Sonora state (Mexico) and the rest in southwestern Arizona and southeastern
California (United States). This is the warmest and the most arid North
American desert due to its low elevation (<600 m) and 22-24 °C average annual
temperature. The bi-seasonal (winter and summer) rainfall averages 200-400
mm per year (Hernandez 2006).

The desert vegetation is characterized by trees and shrubs, including
Parkinsonia florida (A. Gray) S. Watson, Olneya tesota A. Gray, Prosopis
glandulosa Torr., Fouquieria macdougalii Nash, and Bursera spp. The majority
of the cacti are large, with Carnegiea gigantea (Engelm.) Britton & Rose the

146 ... Lizarraga & al.

TABLE 1. Studied Sonoran localities and types of vegetation

LOCALITY LATITUDE N LONGITUDE W ALTITUDE VEGETATION
Town of Benjamin Hill

1. Las Animas 30°12’37.95” 111°18’55.04” 770m MDS
Town of Opodepe

2. Tuape 30°02’52.89” 111°00’22.05” 670m M
Town of Guaymas

3. Cafién de Nacapule 27°59'08.54” 111°02’06.40” 81m SC

4. El Tigre 28°06'31.14” 111°01’21.11” 139m SC

5. El Apache 28°19'25.33” 111°14’27.74” 43m M

6. Maytorena 28°13'32.49” 110°48’50.28” 40m SC
Town of Carbo

7. San Luis 29°33’42.38” 111°04’52.79” 458 m MDS
Town of Hermosillo

8. La Primavera 28°48’06.37” 111°09’09.62” 167 m MDS

9. La Pintada 28°39'56.35” 110°57'15.64” 233 m SC

10. El Papalote 29°12'56.14” 111°02’39.43” 348 m M

Types of vegetation: MDS = microphyllous desert scrub; SC = sarcocaule scrub; M = mezquital

most representative (Hernandez 2006). Myxomycetes can develop in arid
environments due to decomposition of and water retained by succulent plants
such as Opuntia spp. These provide an excellent microhabitat with enough
moisture for optimal development. Although the Sonoran desert has a high
level of biological diversity, here have been only a few studies of its myxobiota.

The Sonoran desert records of Evenson (1961) and Blackwell & Gilbertson
(1980a,b; 1984) were obtained mainly in Arizona (U.S.A.) in the United States.
Although there have been various diverse studies of the myxobiota in Sonora
(Mexico), only twenty-seven reports are known for the biosphere reserve
El Pinacate and Gran Desierto de Altar (Esqueda et al. 2013, Moreno et al.
2004); most studies have been centered in pine-oak forests and lowland areas
(Pérez-Silva et al. 2001; Moreno et al. 2006; Lizarraga et al. 2007, 2008a,b).

The catalogue of myxomycetes for Sonora state (Moreno et al. 2007) has
been increased from 77 species to 108 species by Lizarraga et al. (2007, 2008a,b)
and Esqueda et al. (2013). The present contribution brings the total number to
Lis,

Materials & methods

Four surveys were conducted in 2009, one each season of the year, in 10 localities
of the central plains of the Sonoran desert (TABLE 1). Specimens were gathered or
cultured in moist chamber cultures. Permanent slides were prepared in Hoyer’s liquid
for microscopic study. Collections have been deposited in the collection of fungi of the

Myxomycetes of Sonora 6 (Mexico) ... 147

Universidad Estatal de Sonora, Hermosillo, Sonora, México (UES), with a small part in
the herbarium of the Universidad de Alcala, Alcala de Henares, Madrid, Spain (AH).
The spore ornamentation was examined under SEM using the material deposited in
AH. Collector names are abbreviated as follows: AG = A. Gutiérrez; ML = M. Lizarraga;
CS = C. Salazar; and AS = A. Sanchez. New species records for Sonora are marked with a
single asterisk, and the species recorded as new for Mexico is marked with two asterisks.

Taxonomy

Arcyria cinerea (Bull.) Pers.

STUDIED MATERIAL: MEXICO. Sonora: Loc. 4, on Forchhammeria watsonii Rose,
leg. ML, CS, AS & AG, 19-VIII-2009, obtained from moist chamber, 30-IX-2009 (UES
8539); obtained from moist chamber, 9-X-2009 (UES 8541).

ComMENTS— Previously cited for Sonora by Lizarraga et al. (2007, 2008b).

Arcyria denudata (L.) Wettst.

STUDIED MATERIAL: MEXICO. Sonora: Loc. 1, on decomposing wood, leg. AG & AS,
22-V-2009, obtained from moist chamber, 11-VI-2009 (UES 8106).

CoMMENTS—Previously recorded for Sonora by Pérez-Silva et al. (2001), and
Lizarraga et al. (2008a,b).

Badhamia gracilis (T. Macbr.) T. Macbr.

STUDIED MATERIAL: MEXICO. Sonora: Loc. 1, on Cylindropuntia sp., leg. AS & AG,
9-III-2009, obtained from moist chamber, 20-IV-2009 (UES 7858); Loc. 2, on Opuntia
sp., leg. AS & AG, 14-II-2009, obtained from moist chamber, 15-IV-2009 (UES 7854);
Loc. 3, on Stenocereus thurberi (Engelm.) Buxb., leg. AS & AG, 26-V-2009, obtained
from moist chamber, 30-VI-2009 (UES 8206); Loc. 4, on Lophocereus schottii (Engelm.)
Britton & Rose, leg. CS, AS & AG, 7-XI-2009, obtained from moist chamber, 4-XII-2009
(UES 8660); Loc. 5, on Parkinsonia sp., obtained from moist chamber, 20-IV-2009 (UES
7856); Loc. 6, on Cylindropuntia versicolor (Engelm.) EM. Knuth, leg. AG & AS, 12-
III-2009, obtained from moist chamber, 6-IV-2009 (UES 7845); Loc. 7, on Stenocereus
thurberi, obtained from moist chamber, 14-IV-2009 (UES 7962); Loc. 8, on Lophocereus
schottii, leg. AS & AG, 14-III-2009, obtained from moist chamber, 14-IV-2009 (UES
7847); Loc. 9, on Cylindropuntia arbuscula (Engelm.) EM. Knuth, leg. AG & AS 26-V-
2009, obtained from moist chamber, 11-VI-2009 (UES 8110); Loc. 10, on Parkinsonia
sp. leg. AS & AG, 12-III-2009, obtained from moist chamber, 20-IV-2009 (UES 7860).

COMMENTS— We follow the taxonomy of Moreno & Oltra (2010), who regarded
Badhamia melanospora Speg. as separate from B. gracilis. The two species differ
mainly in spore morphology, with the spores >15-17 um diam., polyhedral,
and densely warty in B. melanospora and <12-14 um diam, polyhedral, and
with scattered warts with intervening spaces in B. gracilis. Both species fruit
mainly on succulent plants (Castillo et al. 1996). Some studied collections were
mistakenly identified as B. melanospora by Moreno et al. (2006) and Esqueda
et al. (2013).

148 ... Lizarraga & al.

Badhamia spinispora (Eliasson & N. Lundg.) H.W. Keller & Schokn. FIGS 1, 5-6
Fructifications sessile, subglobose to short plasmodiocarps, white to
grey, growing grouped together. Sporotheca 0.2-1 x 0.2-0.5 mm. Peridium
double, calcareous, with irregular dehiscence. Hypothallus inconspicuous.
Pseudocolumella irregular, calcareous. Capillitium scarce and joined by
strands consisting of irregular calcareous nodules. Spores (9-)10-12 x 13-15
um, globose to subglobose, black in mass, violaceous with a paler area under
LM, ornamented with large spines. SEM shows that the spore ornamentation is
formed by irregularly scattered or grouped <0.5 um high warts.
STUDIED MATERIAL: MEXICO. Sonora: Loc. 9, over hare manure, leg. AG & AS, 26-V-
2009; obtained from moist chamber, 30-VI-2009 (UES 8116, AH 42974).
ComMENTS— This species is characterized by its generally coprophilous habitat
(although it does occasionally grow on tree bark), cespitose habit, capillitium
that varies from physaroid to badhamioid, and spores that are dark in mass,
spiny, and with a paler area. A study of B. spinispora, Physarum apiculosporum
Hark., and Badhamia verrucospora G. Moreno & al. was published by Moreno
et al. (2011).
Cited from Sonora by Esqueda et al. (2012, 2013).

Comatricha elegans (Racib.) G. Lister

STUDIED MATERIAL: MEXICO. Sonora: Loc. 1, on Olneya tesota, leg. AG & AS, 22-V-

2009, obtained from moist chamber, 3-VI-2009 (UES 8120); Loc. 7, leg. ML, CS, AS &

AG, 17-VIII-2009, obtained from moist chamber, 7-IX-2009 (UES 8563); Loc. 9, 20-

VIII-2009, obtained from moist chamber, 9-IX-2009 (UES 8621); Loc. 10, decomposing

wood, 21-V-2009, obtained from moist chamber, 11-VI-2009 (UES 8122).
CoMMENTS—Lizarraga et al. (2005) published a study (including SEM
photomicrographs) of C. elegans from Chihuahua.

Previously recorded from Sonora by Lizarraga et al. (2007, 2008a).

**Comatricha mirabilis R.K. Benj. & Poitras Figs 7-12

Fructifications stipitate, 0.7-0.9 mm tall. Sporotheca subglobose to
subcylindrical, 0.4-0.6 mm diam. Peridium fleeting and persistent at the
base, resembling a necklace. Stipe short, dark, 0.3-0.6 mm high. Hypothallus
membranous, hyaline, as common in Comatricha. Columella crossing the
sporotheca and forking into several branches at the apex. Capillitium parallel
along the columella, dark brown with free ends, dichotomous, widened and
forked at the tips that protrude at the periphery of the sporotheca. Spores dark
brown in mass, 9-12 um diam., globose to subglobose, reticulate, with walls
1-2 um high. SEM observation reveals that the spore is reticulate and the high
banded reticulum has entire and unperforated walls.

Myxomycetes of Sonora 6 (Mexico) ... 149

Ficures 1-6. Badhamia spinispora (AH 42974): 1. Plasmodiocarps. 5-6. Spores (SEM). Cribraria
confusa (AH 42976): 2-4. Sporocarps. Bars: 1 = 1 mm; 2-4 = 0.5 mm; 5-6 = 2 um.

STUDIED MATERIAL: MEXICO. Sonora: Loc. 4, on bark of Forchhammeria watsonii,

leg. AG & AS, 27-V-2009, obtained from moist chamber, 30-VI-2009 (UES 8129, AH

42975).
CoMMENTS—Comatricha mirabilis is characterized by its capillitium that
extends parallel along the columella without forming a network with bifurcated
ends that protrude from the sporotheca and fully reticulate globose spores. It
was described growing over goat manure in USA in a moist chamber (Benjamin
& Poitras 1950). Later, Mandell & Blackwell (2008) gathered abundant material
from tree barks of Tamarix arabica, Prosopis juliflora, Phoenix dactylifera, and
Cordia myxa from a Bahrain desert in the Kingdom of Bahrain archipelago
near Saudi Arabia, which they developed in a moist chamber. Andrade-Bezerra
et al. (2008) described material developed on Brazilian cottontail (Sylvilagus
brasiliensis L.) dung in a moist chamber, Poulain et al. (2011) gathered it from
France on wood debris, and Schnittler et al. (2013) identified the species from
arid areas in China. We obtained three sporocarps on Forchhammeria watsonii
bark in a moist chamber. This is the first record for Mexico.

150 ... Lizarraga & al.

Comatricha tenerrima (M.A. Curtis) G. Lister

STUDIED MATERIAL: MEXICO. Sonora: Loc. 1, on Olneya tesota, leg. AS & AG, 22-V-
2009, obtained from moist chamber, 15-VI-2009 (UES 8124); Loc. 9, on decomposing
wood, leg. CS, AS & AG, 6-XI-2009, obtained from moist chamber, 11-XII-2009 (UES
8685); Loc. 10, on decomposing wood, leg. AG & AS, 21-V-2009, obtained from moist
chamber, 6-VI-2009 (UES 8130).
CoMMENTS— Lizarraga et al. (2007) published a description with SEM
micrographs of C. tenerrima from Sonora. It has also been cited from other
natural protected areas in Sonora (Lizarraga et al. 2008a,b; Esqueda et al. 2013).

*Cribraria confusa Nann.-Bremek. & Y. Yamam. Fras. 2-4

Fructifications small (0.3-0.9 mm high), stipitate. Sporotheca globose to
subglobose, 0.1-0.2 mm diam. Stipe 0.2-0.7 mm long, dark brown, with debris.
Peridium evanescent, forming a small discoid calyculus at the base beneath
an open network of irregular flattened nodules covered by dictydine granules.
Hypothallus membranous. Spores 7-8 um diam., yellow in mass, light yellow
under LM, globose, with a warty ornamentation.

STUDIED MATERIAL: MEXICO. Sonora: Loc. 3, on palm fibers, leg. ML, CS, AS &
AG, 19-VIII-2009, obtained from moist chamber, 5-I[X-2009 (UES 8570); obtained
from moist chamber, 7-IX-2009 (UES 8759, AH 42976 with Arcyria cinerea and
Paradiacheopsis sp.); on Guaiacum coulteri A. Gray, obtained from moist chamber, 3-X-
2009 (UES 8571).
ComMMENtTS—Cribraria confusa has been recorded only from Chihuahua on
decomposing wood of Pinus sp. by Lizarraga et al. (2003), who presented LM
and SEM micrographs.

Cribraria violacea Rex
STUDIED MATERIAL: MEXICO. Sonora: Loc. 10, on palm fibers, leg. AG, 21-V-2009,
obtained from moist chamber, 15-VI-2009 (UES 8135).
CoMMENTS— Moreno et al. (2001) published an SEM study of the spore
ornamentation on material from Baja California.
Previously cited from Sonora by Moreno et al. (2006) and Lizarraga et al.
(2008b).

Dictydiaethalium plumbeum (Schumach.) Rostaf.

STUDIED MATERIAL: MEXICO. Sonora: Loc. 1, on decomposing wood, leg. ML, CS, AS
& AG, 17-VIII-2009, obtained from moist chamber, 9-IX-2009 (UES 8572); Loc. 5, leg.
AG & AS, 14-III-2009, obtained from moist chambers, 1-V-2009 (UES 7880).

CoMMENTS—Previously cited from Sonora by Moreno et al. (2006) and
Lizarraga et al. (2008b).

Myxomycetes of Sonora 6 (Mexico) ... 151

FIGURES 7-17. Comatricha mirabilis (AH 42975): 7. Sporocarp. 8. Capillitium. 9. Spores.
10. Capillitium endings (detail, SEM). 11-12. Spores by SEM. Didymium eremophilum (AH
42977): 13. Sporocarps. 14. Sporocarp (detail). 15-17. Variability of spore ornamentation (SEM).
Bars: 7 = 0.1 mm; 8 = 50 um; 9 = 10 um; 13-14 = 0.5 mm; 15 = 5 um; 16-17 = 2 um.

152 ... Lizarraga & al.

Didymium dubium Rostaf.

STUDIED MATERIAL: MEXICO. Sonora: Loc. 1, on Olneya tesota, leg. AS & AG,
9-III-2009, obtained from moist chamber, 20-IV-2009 (UES 7888); Loc. 2, on Prosopis
glandulosa, leg. ML, CS, AS & AG, 19-VII-2009, obtained from moist chamber, 9-IX-
2009 (UES 8755); Loc. 5, on Prosopis glandulosa, leg. AS & AG, 14-III-2009, obtained
from moist chamber, 9-IV-2009 (UES 7886); Loc. 6, on Prosopis glandulosa, leg. AG
& AS 12-III-2009, obtained from moist chamber, 25-IV-2009 (UES 7889); Loc. 8, on
Cylindropuntia sp., leg. AG & AS, 14-III-2009, obtained from moist chamber, 14-IV-
2009 (UES 7882); Loc. 9, on Cylindropuntia arbuscula, leg. CS, AS & AG, 6-XI-2009,
obtained from moist chamber, 26-XI-2009 (UES 8692).
ComMMENTS—Moreno et al. (2001) published a SEM study of the spore

ornamentation. Previously cited from Sonora by Moreno et al. (2006).

*Didymium eremophilum M. Blackw. & Gilb. Fics 13-17
Fructifications stipitate, gregarious, 0.2-0.9 mm tall. Sporotheca globose to
subglobose, 0.1-0.3 mm diam., whitish with lighter zones, with a dark disc
at the base. Stipe robust, 0.1-0.5 mm tall, reddish-brown to darkish brown,
wider and darker at the base. Hypothallus membranous, discoid, concolorous
with stipe. Peridium double, with the outer membrane formed by calcareous
crosshairs, the inner membranous with iridescent blue tones. Capillitium and
columella absent. Spores globose to subglobose, black in mass, violaceous
by LM, 9-10 um diam., with a reticulate hemisphere and the opposite warty
(as confirmed by SEM; see Moreno & Mitchell 2013).
STUDIED MATERIAL: MEXICO. Sonora: Loc. 4, leg. CS, AS & AG, 7-IX-2009, obtained
from moist chamber, 4-XII-2009 (UES 8698); Loc. 8, on Lophocereus schottii, leg. AG
& AS, 21-V-2009, obtained from moist chamber, 11-VI-2009 (UES 8141, AH 42977);
obtained from moist chamber, 15-VI-2009 (UES 8142); obtained from moist chamber,
19-VI-2009 (UES 8143); obtained from moist chamber, 22-VI-2009 (UES 8144);
obtained from moist chamber, 30-VI-2009 (UES 8145); leg. CS, AS & AG, 6-XI-2009,
obtained from moist chamber, 30-XI-2009 (UES 8699); obtained from moist chamber,
11-XII-2009 (UES 8700); obtained from moist chamber, 12-XII-2009 (UES 8701);
obtained from moist chamber, 12-XII-2009 (UES 8702); Loc. 9, on cactus remains, leg.
AG & AS, 26-V-2009, obtained from moist chamber, 15-VI-2009 (UES 8146).

ComMMENTS—Didymium eremophilum is a succulenticolous species adapted to
arid environments (Blackwell & Gilbertson 1980b), characterized by its small
size, stipitate sporocarps, crystalline lime, lack of capillitium, partly reticulate
verrucose spores, and succulenticolous habitat (Moreno & Mitchell 2013).

Didymium eremophilum specimens, very difficult to find due to their small
size, have thus far been found only from moist chamber. Previously recorded in
Mexico only from Puebla (Estrada-Torres et al. 2009), our study provides a first
record for the Sonoran mycobiota.

Myxomycetes of Sonora 6 (Mexico) ... 153

Didymium squamulosum (Alb. & Schwein.) Fr.

STUDIED MATERIAL: MEXICO. Sonora: Loc. 4, on deer manure, leg. AG & AS 12-
III-2009, obtained from moist chamber, 1-V-2009 (UES 7881); Loc. 7, on Stenocereus
thurberi, leg. AG & AS, 11-II-2009, obtained from moist chamber, 6-IV-2009 (UES
7890).

CoMMENTS— Lizarraga et al. (1999a) published a SEM study of the spore
ornamentation. Widely cited for Mexico (Moreno et al. 2007), D. squamulosum

has been previously cited for Sonora by Moreno et al. (2006) and Lizarraga et
al. (2008b).

Didymium sturgisii Hagelst. Figs 18-23

STUDIED MATERIAL: MEXICO. Sonora: Loc. 7, on Prosopis glandulosa, leg. AG & AS,
11-III-2009, obtained from moist chamber, 8-IV-2009 (UES 7884, AH 42978).

ComMMENTS—This species is characterized by its dirty white to whitish grey
flattened to subglobose fructifications (occasionally with small superficial
orifices) and sporocarp interiors with erect pillars or internally traversing
trabeculae. Moreno et al. (2001) published an SEM study of the capillitium,
pillars, and spores.

Didymium sturgisii has been recorded from Baja California (Moreno et al.
2007), Chihuahua (Salazar-Marquez et al. 2014), Puebla (Estrada-Torres et al.
2009), and Sonora (Esqueda et al. 2012, 2013).

Echinostelium apitectum K.D. Whitney

STUDIED MATERIAL: MEXICO. Sonora: Loc. 1, on Prosopis glandulosa leg. AS & AG,
9-III-2009, observed in moist chamber, 28-III-2009, 4-IV-2009 (UES 7892); Loc. 3, on
Stenocereus thurberi, 19-VIII-2009, obtained in moist chamber, 2-IX-2009, 5-IX-2009
(UES 8589); Loc. 4, on Olneya tesota, leg. AG & AS, 3-VI-2009, obtained from moist
chamber, 17-VI-2009 (UES 8149); Loc. 5, on decomposing wood, leg. ML, CS, AS &
AG, 18-VIII-2009, obtained in moist chamber, 2-IX-2009, 5-IX-2009 (UES 8581); Loc.
6, on Prosopis glandulosa, leg. AG & AS, 12-III-2009, observed in moist chamber, 28-
III-2009, 2-IV-2009 (UES 7902); Loc. 7, on Prosopis glandulosa, leg. AG & AS, 22-V-
2009, observed in moist chamber, 3-VI-2009, 22-VI-2009 (UES 8151); Loc. 9, Carnegiea
gigantea, leg. ML, CS, AS & AG, 20-VIII-2009, observed in moist chamber, 2-IX-2009,
7-IX-2009 (UES 8584); Loc. 10, on Prosopis glandulosa, leg. CS, AS & AG, 3-XI-2009,
observed in moist chamber, 9-XI-2009, 30-XI-2009 (UES 8703).

CoMMENTS—Previously cited from Sonora by Lizarraga et al. (2007, 2008a).

Echinostelium arboreum H.W. Keller & T.E. Brooks

STUDIED MATERIAL: MEXICO. Sonora: Loc. 1, on decomposing wood, leg. AG & AS,
22-V-2009, obtained from moist chamber, 11-VI-2009 (UES 8106 with Arcyria cinerea).

154 ... Lizarraga & al.

ComMENtTS—Moreno et al. (2001) published a SEM study of the sporocarp and
spores. Previously cited from Sonora by Esqueda et al. (2013), E. arboreum has
been widely reported from Mexico (Moreno et al. 2007).

Echinostelium colliculosum K.D. Whitney & H.W. Keller

STUDIED MATERIAL: MEXICO. Sonora: Loc. 1, on Olneya tesota, leg. AS & AG,
9-III-2009, 28-III-2009, obtained from moist chamber, 4-I[V-2009 (UES 7891); Loc.
5, on Guaiacum coulteri, 14-III-2009, obtained from moist chamber, 4-IV-2009 (UES
7893); on Olneya tesota, observed in moist chamber, 4-IV-2009 (UES 7894); on Prosopis
glandulosa, obtained from moist chamber, 8-IV-2009 (UES 7895); on Parkinsonia sp.,
leg. AG & AS, 26-V-2009, obtained from moist chamber, 11-VI-2009 (UES 8152); Loc.
6, on Cylindropuntia arbuscula, leg. AS & AG, 27-V-2009, 3-VI-2009, obtained from
moist chamber, 11-VI-2009 (UES 8153); on Parkinsonia sp., leg. ML, CS, AS & AG, 19-
VIII-2009, in moist chamber, 2-IX-2009, 5-IX-2009 (UES 8588); Loc. 7, on Parkinsonia
sp., leg. AS & AG, 11-III-2009, in moist chamber, 9-IV-2009 (UES 7901); on Encelia
farinosa A. Gray ex Torr., 22-V-2009, 3-VI-2009, in moist chamber, 11-V1I-2009 (UES
8150); on decomposing wood, leg. ML, CS, AS & AG, 17-VIII-2009, in moist chamber,
2-IX-2009, 5-IX-2009 (UES 8591); Loc. 9, on Guaiacum coulteri, 11-III-2009, 28-III-
2009, in moist chamber, 4-IV-2009 (UES 7896); on Olneya tesota, in moist chamber,
4-IV-2009 (UES 7898); on Cylindropuntia arbuscula, in moist chamber, 4-IV-2009 (UES
7899); leg. ML, CS, AS & AG, 20-VIII-2009, in moist chamber, 2-IX-2009, 7-IX-2009
(UES 8583); Loc. 10, on Cylindropuntia sp., leg. AS & AG, 12-III-2009, 4-IV-2009 (UES
7900); leg. ML, CS, AS & AG, 18-VIII-2009, in moist chamber, 9-IX-2009 (UES 8582);
Loc. 10, on Cylindropuntia sp., leg. AS & AG, 12-III-2009, 4-IV-2009 (UES 7900).

ComMMENTS— his species is characterized by its (60-)75-90 um high stipitate
sporocarps, globose 9-13 um diam. pale yellow spores that have a spine-like
ornamentation and articular spore wall. A LM study of sporocarps and spores
by Lizarraga et al. (2003) and SEM study by Moreno et al. (2001) have been
published.

Echinostelium colliculosum has been recorded for Baja California Sur and
Chihuahua (Moreno et al. 2007), Oaxaca and Puebla (Estrada-Torres et al.
2009), and Sonora (Esqueda et al. 2013).

Echinostelium minutum de Bary

STUDIED MATERIAL: MEXICO. Sonora: Loc. 1, on Prosopis glandulosa, leg. ML, CS, AS
& AG, 17-VIII-2009, in moist chamber, 2-IX-2009, 7-IX-2009 (UES 8580); Loc. 3, on
Stenocereus thurberi, leg. ML, CS, AS & AG, 17-VHI-2009, 7-IX-2009 (UES 8590); Loc. 7,
on Prosopis glandulosa, leg. AG, 22-05-2009, obtained from moist chamber, 11-06-2009

FIGURES 18-27. Didymium sturgisii (AH 42978): 18. Fructifications flattened with some calcium
carbonate granules. 19. Sporocarp (detail of small superficial orifices). 20. Capillitium and
erect pillars. 21-22. Spores (SEM). 23. Spore ornamentation (detail, SEM). Licea denudescens
(AH 42979): 24. Sporocarps. 25. Spore (SEM). Licea succulenticola (AH 42980): 26. Sporocarps.
27. Spore (SEM). Bars: 18-19 = 0.5 mm; 20 = 0.25 mm; 21-22 = 2 um; 23 = 1 um.

Myxomycetes of Sonora 6 (Mexico) ... 155

156 ... Lizarraga & al.

(UES 8154); Loc. 9, on Carnegiea gigantea, 9-I[X-2009 (UES 8585); Loc. 10, on Prosopis
glandulosa, leg. CS, AS & AG, 3-XI-2009, in moist chamber, 9-XI-2009, II-2009 (UES
8704).
ComMMENTS— Moreno et al. (2001) published a SEM study of the sporocarp
and spores. Previously reported from Sonora by Moreno et al. (2006).

Enerthenema papillatum (Pers.) Rostaf.

STUDIED MATERIAL: MEXICO. Sonora: Loc. 4, on Forchhammeria watsonii, leg. ML,

CS, AS & AG, 19-VIII-2009, obtained in moist chamber, 5-IX-2009 (UES 8758).
CoMMENTS— Lizarraga et al. (1999a) published a SEM study of the spores from
collections from Baja California, Mexico. Widely recorded in Mexico (Moreno
et al. 2007), E. papillatus was recorded previously from Sonora by Moreno et
al. (2006).

Fuligo cinerea (Schwein.) Morgan
STUDIED MATERIAL: MEXICO. Sonora: Loc.10, on sheep manure, leg., AG, 21-V-2009,
obtained from moist chamber, 30-VI-20009 (UES 8118).
COMMENTS—Sometimes similar in size as some Badhamia species (e.g.,
B. spinispors, B. verrucospora G. Moreno et al.), Fuligo cinerea is separated by
its ovoid spores covered by small warts that join together to form a reticulum.
Cited previously for Sonora by Braun & Keller (1986) and Lizarraga et al.
(2007).

Fuligo septica (L.) EH. Wigg.

STUDIED MATERIAL: MEXICO. Sonora: Loc. 9, on soil, leg. ML, 11-II-2009 (UES
8677).

CoMMENTS— Lizarraga et al. (1999a) published a SEM study of the spores.
Fuligo septica has a wide distribution in Mexico (Moreno et al. 2007) and has
been cited for Sonora by Pérez-Silva et al. (2001) and Lizarraga et al. (2007,
2008a,b).

Licea biforis Morgan

STUDIED MATERIAL: MEXICO. Sonora: Loc. 1, on Olneya tesota, leg. AG & AS, 9-III-
2009, obtained in moist chamber, 6-IV-2009 (UES 7905); Loc. 2, on Prosopis glandulosa,
leg. AS & AG, 14-III-2009, obtained in moist chamber, 13-IV-2009 (UES 7908); Loc. 3,
on Acacia sp., 26-V-2009, obtained in moist chamber, 15-VI-2009 (UES 8159); Loc. 5,
on Guaiacum coulteri bark, leg. AS & AG 14-III-2009, obtained in moist chamber, 20-
IV-2009 (UES 7913); Loc. 6, on Prosopis glandulosa, leg. AG & AS, 27-V-2009, obtained
in moist chamber, 15-VI-2009 (UES 8162); Loc. 9, on Olneya tesota, leg. AG & AS, 11-
III-2009, obtained in moist chamber, 1-V-2009 (UES 7915); Loc. 10, leg. CS, AS & AG,
3-XI-2009, obtained in moist chamber, 12-XII-2009 (UES 8716).

Myxomycetes of Sonora 6 (Mexico) ... 157

CoMMENTS— Lizarraga et al. (1998, 1999b) published SEM studies of the
spores. Widely distributed in Mexico (Moreno et al. 2007), Licea biforis was
previously cited from Sonora by Moreno et al. (2006).

Licea denudescens H.W. Keller & T.E. Brooks Fics 24-25
Fructifications sessile, globose to subglobose, 0.1-0.3 mm diam., 0.1 mm
height, brown to blackish. Peridium membranous with some iridescent tones,
internally ornamented with small buds. Spores violaceous in mass, yellowish
brown by LM, 10-11 um diam., globose to subglobose, walls thick with pale
(thinner) areas, smooth by SEM.
STUDIED MATERIAL: MEXICO. Sonora: Loc. 1, on Olneya tesota, leg. AS & AG, 9-III-
2009, obtained in moist chamber, 8-IV-2009 (UES 7916, AH 42979); Loc. 5, on Olneya
tesota, leg. CS, AS & AG, 6-XI-2009, obtained in moist chamber, 12-XII-2009 (UES
8711); Loc. 7, on Parkinsonia sp., 11-III-2009, obtained in moist chamber, 20-IV-2009
(UES 7918); obtained in moist chamber, 1-V-2009 (UES 7917); Loc. 8, on Acacia greggii
A. Gray, leg. ML, CS, AS & AG, 18-VIII-2009, obtained in moist chamber, 3-X-2009
(UES 8605); Loc. 9, on cactus remains, 11-III-2009, observed in moist chamber, 15-IV-
2009 (UES 7919).
ComMMENTS—Licea denudescens is diagnosed by its thick-walled spores with
thinner areas and papillae found on the inside of the peridium. It has been
cited in Mexico from Puebla (Estrada-Torres et al. 2009), Chihuahua (Salazar-
Marquez et al. 2014), and Sonora (Esqueda et al. 2013).

Licea kleistobolus G.W. Martin

STUDIED MATERIAL: MEXICO. Sonora: Loc. 8, on Acacia greggii bark, leg. ML, CS, AS

& AG, 18-VIII-2009, observed in moist chamber, 7-IX-2009 (UES 8608).
ComMENTS—Moreno et al. (2001) published a SEM study of Licea kleistobolus,
which is characterized by its circular to subglobose fructifications with a
characteristic operculum and warted spores with groups of denser warts. Cited
previously from Sonora by Moreno et al. (2006) and Lizarraga et al. (2007).

Licea succulenticola Mosquera, Lado, Estrada & Beltran-Tej. Fics 26-27

Fructifications sessile, yellowish to dark-yellow at mature, dispersed to
gregarious, subglobose, oval to elongated, 0.1-0.3 mm long, 0.1-0.2 mm
diam., and 0.1 mm tall, with an apical and longitudinal dehiscence. Peridium
membranous, ornamented on its inner part with small warts. Spores yellow in
mass, light yellow by LM, globose to subglobose, (11-)12-14 um diam., with
a lightly paler area, with a slightly warty ornamentation. SEM examination
shows that the spore ornamentation is formed by small croziers more or less
uniformly distributed over the in spore surface.

STUDIED MATERIAL: MEXICO. Sonora: Loc. 2, on Opuntia sp., 5-XI-2009, observed in
moist chamber, 14-XII-2009 (UES 8712); Loc. 3, on Cylindropuntia sp., leg. AS & AG,

158 ... Lizarraga & al.

14-III-2009, obtained in moist chamber, 28-III-2009, 20-IV-2009 (UES 7924); obtained
in moist chamber, 25-IV-2009 (UES 7925, 7926); Loc. 4, on Lophocereus schottii, leg.
AS & AG, 12-III-2009, obtained from moist chamber, 20-IV-2009 (UES 7923); Loc.
6, on Lophocereus schottii, leg. CS, AS & AG, 7-XI-2009, observed in moist chamber,
9-X1-2009, 9-XII-2009 (UES 8726); Loc. 7, on Lophocereus schottii, leg. AS & AG, 11-
III-2009, observed in moist chamber, 28-III-2009, 20-IV-2009 (UES 7927); obtained in
moist chamber, 1-V-2009 (UES 7928, 7929); on Stenocereus thurberi, leg. ML, CS, AS &
AG, 17-VIII-2009, observed in moist chamber, 2-IX-2009, 7-IX-2009 (UES 8747, AH
42980); obtained in moist chamber, 14-IX-2009 (UES 8602); obtained in moist chamber,
21-IX-2009 (UES 8604); obtained in moist chamber, 21-[X-2009 (UES 8603); Loc. 8, on
Lophocereus schottii, leg. ML, CS, AS & AG, 18-VIII-2009, observed in moist chamber,
2-IX-2009, 9-IX-2009 (UES 8594); obtained in moist chamber, 14-IX-2009 (UES 8595);
obtained in moist chamber, 24-IX-2009 (UES 8596); obtained in moist chamber, 30-
IX-2009 (UES 8597); obtained in moist chamber, 5-X-2009 (UES 8598, AH 42981); on
Stenocereus thurberi, leg. CS, AS & AG, 6-XI-2009, observed in moist chamber, 9 -XI-
2009, 14-XII-2009 (UES 8725); Loc. 10, on Cylindropuntia arbuscula, leg. AG & AS,
12-III-2009, obtained in moist chamber, 20-IV-2009 (UES 7920); on cactus remains,
obtained in moist chamber, 20-IV-2009 (UES 7921); obtained in moist chamber, 1-V-
2009 (UES 7922); on Lophocereus schottii, leg. ML, CS, AS & AG, 18-VIII-2009, obtained
in moist chamber, 2-IX-2009, 17-IX-2009 (UES 8599); obtained in moist chamber, 30-
IX-2009 (UES 8600); obtained in moist chamber, 30-IX-2009 (UES 8601).

ComMMENTS—Licea succulenticola resembles L. pumila G.W. Martin & R.M.
Allen (Martin & Alexopoulos 1969) in sporothecal form and spore colour.
(see Mosquera et al. 2003 for differences). Previously recorded from Hidalgo,
Morelos, Puebla, and Tlaxcala (Mosquera et al. 2003), Puebla (Estrada-Torres
et al. 2009), Sonora (Esqueda et al. 2013), and Chihuahua (Salazar-Marquez et
al. 2014).

Paradiacheopsis fimbriata (G. Lister & Cran) Hertel ex Nann.-Bremek. Fics 28-30

STUDIED MATERIAL: MEXICO. Sonora: Loc. 3, on Cylindropuntia sp., leg. AG,
14-III-2009, observed in moist chamber, 28-III-2009, 13-IV-2009 (UES 7943,
AH 42982).
COMMENTS— Widely distributed in Mexico (Moreno et al. 2007), P. fimbriata
was reported from Sonora by Moreno et al. (2006).

Perichaena depressa Lib.

STUDIED MATERIAL: MEXICO. Sonora: Loc. 1, on rabbit manure, leg. ML, CS, AS &

AG, 17-VIII-2009, obtained in moist chamber, 9-X-2009 (UES 8628); Loc. 9, on horse

manure, 20-VIII-2009, obtained in moist chamber, 5-X-2009 (UES 8629).
CoMMENTS— Lizarraga et al. (1998) and Moreno et al. (2001) published SEM
studies of spores and capillitium, comparing type material with samples from
Baja California and Sinaloa. Widely distributed throughout Mexico (Moreno
et al. 2007), P. depressa was previously cited for Sonora by Moreno et al. (2006).

Myxomycetes of Sonora 6 (Mexico) ... 159

FIGURES 28-36. Paradiacheopsis fimbriata (AH 42982): 28. Sporocarp. 29. Sporotheca (detail).
30. Fiber stalk (detail). Perichaena stipitata (AH 42983): 31-32. Stipitate sporocarp (detail).
33. Peridial inner surface (detail). 34. Capillitium with irregular outline. 35-36. Spores (SEM).
Bars: 28-29 = 0.1 mm; 30 = 0.02 mm; 31-32 = 0.5 mm; 33, 35-36 = 1 um; 34 = 5 um.

*Perichaena luteola (Kowalski) Gilert

STUDIED MATERIAL: MEXICO. Sonora: Loc. 10, on cow manure, leg. ML, CS, AS &

AG, 18-VIII-2009, obtained in moist chamber, 27-IX-2009 (UES 8749); 3-X-2009 (UES

8627).
ComMMENTS—The above collections have 10-13 um diam. globose to
subglobose spores that are yellow in mass (lighter yellow by LM) and with a
verruculose episporium and a yellow 2-3um diam. capillitium with smooth
surface. Lizarraga et al. (1999b) published a SEM study of material from Baja
California. Perichaena luteola is widely distributed in Mexico (Moreno et al.
2007).

160 ... Lizarraga & al.

*Perichaena stipitata Lado, Estrada & D. Wrigley Figs 31-36
Fructifications short-stipitate, solitary to gregarious, 0.3-0.8 mm tall. Stipe
robust, cylindrical, black, 0.1-0.2 mm long. Sporotheca globose to subglobose,
rarely obpyriform, yellow, 0.2-0.5 mm diam. Peridium brownish on the base,
inner surface ornamented with small rings by LM and SEM. Hypothallus dark,
membranous and irregular. Capillitium sparse, formed by yellowish branched
filaments of approximately 2 um diam., flexible, of irregular outline. Spores
yellow in mass, light yellow by LM, 12-15 um diam., globose, warted. SEM
examination shows that the spore is ornamented by warts with smooth apices
and which sometimes join into small groups.
STUDIED MATERIAL: MEXICO. Sonora: Loc. 9, on cactus remains, leg. AG & AS, 26-
V-2209, obtained in moist chamber, 30-VI-2009 (UES 8210, AH 42983); on Lophocereus
schottii, leg. ML, CS, AS & AG, 20-VIII-2009, obtained in moist chamber, 17-IX-2009
(UES 8762); obtained in moist chamber, 24-IX-2009 (UES 8761); obtained in moist
chamber, 3-X-2009 (UES 8760, AH 42984); obtained in moist chamber, 9-X-2009 (UES
8643); Loc. 10, on Lophocereus schottii, leg. ML, CS, AS & AG, 18-VII-2009, obtained
in moist chamber, 5-X-2009 (UES 8642).

COMMENTS—Stipitate fructifications and an inner peridium ornamented by
small rings characterize this succulenticolous species. First described as new
to science by Estrada-Torres et al. (2009) from Puebla and Queretaro, Mexico,
P stipitata was later described from Arizona (USA) by Moreno & Mitchell (2013),
who published a colour plate showing its development and morphological
characteristics. A new record for Sonora.

Perichaena vermicularis (Schwein.) Rostaf.

STUDIED MATERIAL: MEXICO. Sonora: Loc. 1, on Carnegiea gigantea, leg. CS, AS
& AG, 5-XI-2009, obtained in moist chamber, 11-XII-2009 (UES 8734); Loc. 2, on
Cylindropuntia arbuscula, leg. AS & AG, 22-V-2009, obtained in moist chamber, 25-VI-
2009 (UES 8191); Loc. 3, on Cylindropuntia sp., leg. AS & AG 14-III-2009, obtained in
moist chamber, 15-IV-2009 (UES 7953); Loc. 4, on Forchhammeria watsonii, leg. AG &
AS, 12-III-2009, obtained in moist chamber, 3-IV-2009 (UES 7944); Loc. 6, Parkinsonia
sp., 19-VIH-2009, obtained in moist chamber, 24-IX-2009 (UES 8633); Loc. 7, on
Cactaceae remains, 11-III-2009, obtained in moist chamber, 8-IV-2009 (UES 7947);
obtained in moist chamber, 20-IV-2009 (UES 7948); Loc. 8, on Cactaceae remains,
obtained in moist chamber, 18-VIII-2009, 17-IX-2009 (UES 8631).

ComMENtTs—Moreno et al. (2001) published a SEM study of spores and
capillitium in collections from Baja California that grew cespitose in moist
chamber over different substrates. Widely distributed in Mexico (Moreno et
al. 2007), P. vermicularis was previously reported from Sonora by Moreno et
al. (2006).

Myxomycetes of Sonora 6 (Mexico) ... 161

Physarum album (Bull.) Chevall.

STUDIED MATERIAL: MEXICO. Sonora: Loc. 7, on Prosopis glandulosa, leg. AG & AS,
17-VIUI-2009, obtained in moist chamber, 9-IX-2009 (UES 8651); Loc. 10, on Prosopis
glandulosa, leg. ML, CS, AS & AG, 18-VIII-2009, obtained in moist chamber, 7-IX-2009
(UES 8650).
CoMMENTS—Lizarraga et al. (1999b) published a SEM study of spore
ornamentation in collections from Baja California. Widely distributed in
Mexico (Moreno et al. 2007) and previously reported for Sonora (Lizarraga
et al. 2007, 2008b), Physarum album frequently grows on many different
substrates when cultivated in moist chamber.

Physarum decipiens M.A. Curtis
Fructifications sessile, dispersed to grouped, subglobose or forming
short plasmodiocarp. Sporotheca yellow-greenish, 0.3-1.2 x 0.2-0.6 mm,
membranous, peridium encrusted with calcium carbonate. Capillitium
physaroid. Spores 11-13 um diam., subglobose to globose, violaceous and
slightly warty.
STUDIED MATERIAL: MEXICO. Sonora: Loc. 1, on Prosopis glandulosa, leg. AG & AS,
9-III-2009, obtained in moist chamber, 6-IV-2009 (UES 7967); 22-V-2009, obtained in
moist chamber, 11-VI-2009 (UES 8193); Loc. 2, on Prosopis glandulosa, leg. AS & AG 14-
III-2009, obtained in moist chamber, 8-IV-2009 (UES 7970); Loc. 4, on Forchhammeria
watsonii, leg. ML, CS, AS & AG, 19-VIII-2009, obtained in moist chamber, 7-IX-2009
(UES 8649).
ComMENtTsS—Morphologically, P decipiens is highly variable. Previously cited
from Chihuahua (Lizarraga et al. 2003), Oaxaca and Puebla (Estrada-Torres et
al. 2009), Tlaxcala (Rodriguez-Palma et al. 2002), and recently Sonora (Esqueda
et al. 2012, 2013).

Physarum leucophaeum Fr.

STUDIED MATERIAL: MEXICO. Sonora: Loc. 1, on Olneya tesota, leg. AS & AG, 22-V-
2009, obtained in moist chamber, 25-VI-2009 (UES 8194); Loc. 5, on Parkinsonia sp.,
leg. AS & AG 14-III-2009, obtained in moist chamber, 1-V-2009 (UES 7966); Loc. 6, on
Encelia farinosa, 27-V-2009, obtained in moist chamber, 22-VI-2009 (UES 8198); Loc.
8, on Larrea tridentata Coville, leg. AS & AG, 21-V-2009, obtained in moist chamber,
30-VI-2009 (UES 8202); Loc. 10, on Guaiacum coulteri, leg. ML, CS, AS & AG, 18-VIII-
2009, obtained in moist chamber, 9-X-2009 (UES 8653).

ComMMENTS— Moreno et al. (2001) published a SEM study of spore
ornamentation in collections from Baja California. The recently named
species from arid regions of Eurasia and North and South America, Physarum
pseudonotabile Novozh. et al. (Novozhilov et al. 2013), is very close to P. notabile

162 ... Lizarraga & al.

T. Macbr. and P leucophaeum; distinguishing the three species is complex
and additional ecological, morphological, and molecular studies are needed.
Physarum leucophaeum is widely distributed in Mexico (Moreno et al. 2007),
common when cultured in moist chamber. Previously cited from Sonora by
Lizarraga et al. (2007, 2008b).

Acknowledgments

This research was made possible by support from the following projects: CONABIO
(Proyecto GT016). We wish to express our gratitude to Dr. S.L. Stephenson (University
of Arkansas, U.S.A.) and Mr. D.W. Mitchell (Upper Hartfield, U.K.) for reviewing the
manuscript and providing useful comments. We also extend our thanks to Mr. A. Priego
and Mr. J.A. Pérez (Electron Microscopy Service, University of Alcala de Henares)
for their invaluable help with the SEM. We also thank Luis Monje (Department of
Drawing and Scientific Photography, University of Alcala de Henares) for his help in
digitally preparing the photographs, and we are grateful to Dr. J. Rejos, curator of the
AH herbarium for his assistance with the specimens examined in the present study.
M. Lizarraga extends his gratitude to Dr. Alejandro Martinez Martinez (Jefe del Dpto.
de Ciencias Quimico Bioldgicas, ICB-UACJ) for his assistance in obtaining financial
support. C. Salazar thanks CONACYT for her scholarship to obtain a master’s degree.

Literature Cited

Andrade-Bezerra MF, Teofilo da Silva WM, Cavalcanti LH. 2008. Coprophilous myxomycetes of
Brazil: first report. Revista Mexicana de Micologia 27: 29-37.

Benjamin RK, Poitras AW. 1950. An addition to the myxomycetes genus Comatricha. Mycotaxon
42: 514-518.

Blackwell M, Gilbertson RL. 1980a. Sonoran desert Myxomycetes. Mycotaxon 11:139-149.

Blackwell M, Gilbertson RL. 1980b. Didymium eremophilum: a new myxomycete from the Sonoran
Desert. Mycologia 72: 791-797. http://dx.doi.org/10.2307/3759772

Blackwell M, Gilbertson RL. 1984. Distribution and sporulation phenology of myxomycetes in the
Sonoran desert of Arizona. Microbial Ecology 10: 369-377.
http://dx.doi.org/10.1007/BF02015561

Braun KL, Keller HW. 1986. Myxomycetes of Mexico III. Revista Mexicana de Micologia 2: 25-39.

Castillo A, Illana C, Moreno G. 1996. Badhamia melanospora Speg. A species wrongly interpreted.
Mycotaxon 57: 163-170.

Esqueda M, Lizarraga M, Gutiérrez A, Coronado ML, Valenzuela R, Raymundo T, Chacon §,
Vargas G, Barredo-Pool F. 2012. Diversidad fungica en planicies del desierto central Sonorense
y centro del desierto Chihuahuense. CIAD-UACJ-CESUES-IPN-INECOL-CICY. Informe
Final SNIB-CONABIO Proyecto GT016. México, D.F.

Esqueda M, Coronado ML, Gutiérrez A, Lizarraga M, Raymundo T, Valenzuela R. 2013. Hongos
de Reserva de la Biosfera El Pinacate y Gran Desierto de Altar. Centro de Investigacion en
Alimentacion y Desarrollo, A.C. México. 106 p.

Estrada-Torres A, Wrigley De Basanta D, Conde E, Lado C. 2009. Myxomycetes associated with
dryland ecosystems of the Tehuacan-Cuicatlan Valley Biosphere Reserve, Mexico. Fungal
Diversity 36: 17-56.

Evenson AE. 1961. A preliminary report of the Myxomycetes of Southern Arizona. Mycologia 53:
137-144. http://dx.doi.org/10.2307/3756232

Myxomycetes of Sonora 6 (Mexico) ... 163

Hernandez HM. 2006. La vida en los desiertos mexicanos. Fondo de la Cultura Econdémica,
México, DF. 188 p.

Lizarraga M, Moreno G, Illana C. 1997. The Myxomycetes from Baja California Mexico). I.
Mycotaxon 63: 287-300.

Lizarraga M, Illana C, Moreno G. 1998. First records of Myxomycetes in the state of Sinaloa, Mexico.
Micologia e Vegetazione Mediterranea 13: 167-176.

Lizarraga M, Illana C, Moreno G. 1999a. SEM studies of the Myxomycetes from the Peninsula of
Baja California Mexico), I. Arcyria to Fuligo. Annales Botanici Fennici 35: 287-306.

Lizarraga M, Illana C, Moreno G. 1999b. SEM studies of the Myxomycetes from the Peninsula of
Baja California Mexico), II. Hemitrichia to Trichia. Annales Botanici Fennici 36: 187-210.
Lizarraga M, Moreno G, Singer H, Ilana C. 2003. Myxomycetes from Chihuahua, México.

Mycotaxon 88: 409-424.

Lizarraga M, Moreno G, Illana C, Singer H. 2005. Myxomycetes from Chihuahua, Mexico III.
Mycotaxon 93: 75-88.

Lizarraga M, Moreno G, Esqueda M, Sanchez A, Coronado M. 2007. Myxomycetes from Sonora
Mexico. 3: National Forest Reserve and Wildlife Refuge Ajos-Bavispe. Mycotaxon 99: 291-301.

Lizarraga M, Moreno G, Esqueda M, Coronado M. 2008a. Myxomycetes of Sonora, México. 4:
Sierra de Alamos-Rio Cuchujaqui Biosphere Reserve. Mycotaxon. 103: 153-170.

Lizarraga M, Moreno G, Esqueda M, Coronado M. 2008b. Myxomycetes of Sonora, México. 5: Ajos-
Bavispe National Forest Reserve and Wildlife Refuge and Sierra de Alamos-Rio Cuchujaqui
Biosphere Reserve. Mycotaxon 104: 423-443.

Mandell QA, Blackwell M. 2008. Rare or rarely collected? Comatricha mirabilis from the desert of
Bahrain. Mycologia 100(5): 742-745. http://dx.doi.org/10.3852/07-154

Martin GW, Alexopoulos CJ. 1969. The Myxomycetes. University of Iowa Press. 561 pp.

Moreno G, Mitchell DW. 2013. Three rare species of Myxomycetes from Arizona and New Mexico,
USA. Boletin Sociedad Micolégica de Madrid 37: 77-83.

Moreno G, Oltra M. 2010. Notes on the genus Badhamia, Badhamiopsis and Craterium
(Myxomycetes) in Spain. Boletin Sociedad Micoldgica de Madrid 34: 161-197.

Moreno G, IIlana C, Lizarraga M. 2001. SEM studies of the Myxomycetes from the Peninsula of Baja
California (Mexico), III. Additions. Annales Botanici Fennici 38: 225-247.

Moreno G, Illana C, Esqueda M, Castillo A, Pérez-Silva E. 2004. Notes on Myxomycetes from
México. II: Boletin Sociedad Micoldégica de Madrid 28. 55-63.

Moreno G, Lizarraga M, Esqueda M, Pérez-Silva E, Herrera T. 2006. Myxomycetes de Sonora
México. II: Reserva Forestal Nacional y Refugio de Fauna Silvestre Ajos-Bavispe. Revista
Mexicana de Micologia 22: 13-23.

Moreno G, Lizarraga M, Illana C. 2007. Catalogo de los Myxomycetes de México. Boletin Sociedad
Micoldgica de Madrid 31: 187-229.

Moreno G, Mitchell DW, Novozhilov YK. 2011. A new species of Badhamia (Myxomycetes)
confused with other species with similar morphology. Boletin Sociedad Micoldgica de Madrid
35395. 102:

Mosquera J, Lado C, Estrada-Torres A, Beltran-Tejera E, Wrigley De Basanta D. 2003. Description
and culture of a new myxomycete, Licea succulenticola. Anales Jardin Botanico de Madrid
60(1): 3-10.

Novozhilov YK, Okun MV, Erastova DA, Shchepin ON, Zemlyanskaya IV, Garcia-Carvajal E,
Schnittler M. 2013. Description, culture and phylogenetic position of a new xerotolerant
species of Physarum. Mycologia 105(6): 1535-1546. http://dx.doi.org/10.3852/12-284

164 ... Lizarraga & al.

Pérez-Silva E, Herrera T, Esqueda M, Illana C, Moreno G. 2001. Myxomycetes de Sonora. I.
Mycotaxon 77: 181-192.

Poulain M, Meyer M, Bozonnet J. 2011. Les myxomycétes tomes 1-2. Fédération mycologique et
botanique Dauphiné-Savoie, Sevrier, France.

Rodriguez-Palma M, Varela-Garcia A, Lado C. 2002. Corticolous Myxomycetes associated with
four tree species in México. Mycotaxon 81: 345-355.

Salazar-Marquez C, Lizarraga M, Moreno G. 2014. Myxomycetes de matorral xer6filo del municipio
de Juarez, Chihuahua, México. Boletin Sociedad Micolégica de Madrid 38: 67-77.

Schnittler M, Novozhilov YK, Carvajal E, Spiegel FW. 2013. Myxomycete diversity in the
Tarim basin and eastern Tian-Shan, Xinjiang Prov., China. Fungal Diversity 59: 91-108.
http://dx.doi.org/10.1007/s13225-012-0186-5

MY COTAXON

http://dx.doi.org/10.5248/130.165
Volume 130, pp. 165-171 January-March 2015

New taxa of Ambomucor (Mucorales, Mucoromycotina)
from China

XIAO- YONG Lru* & RU- YONG ZHENG

State Key Laboratory of Mycology, Institute of Microbiology, Chinese Academy of Sciences, Beijing
100101, China

* CORRESPONDENCE TO: liuxiaoyong@im.ac.cn

ABSTRACT — A new species and a new variety, A. ovalisporus and A. seriatoinflatus var.
longior, are described from soils in northwest China. A key to all known taxa of Ambomucor
accompanies descriptions and line drawings of the two new taxa. The maximum growth
temperature used as an auxiliary criterion in classification of Mucorales is also discussed.

KEY worps — morphology, taxonomy, Mucoraceae, Zygomycota

Introduction

The genus Ambomucor (Mucorales), typified by A. seriatoinflatus R.Y.
Zheng & X.Y. Liu, was introduced recently (Zheng & Liu 2014), together
with A. seriatoinflatus var. brevior R.Y. Zheng & X.Y. Liu and A. clavatus
R.Y. Zheng & X.Y. Liu. These taxa were found in northwest China. Recently,
two new Ambomucor taxa were found in the northwest regions (Tibet and
Shaanxi Province) of the country and are described here as A. ovalisporus
and A. seriatoinflatus var. longior. All Ambomucor species and varieties are
characterized by simultaneously having two kinds of sporangia: 1) fertile
sporangia containing a columella and sporangiospores and 2) aborted
sporangia that lack spores and a columella and which proliferate in chains. The
criteria for differentiating the taxa of the enlarged Ambomucor are almost the
same as previously reported (Zheng & Liu 2014). A key to all Ambomucor taxa
is presented.

Materials & methods

Isolation
A small piece of moss, humus, or dung was soaked and stirred with sterilized distilled
water. Different dilutions were poured with PDA media to make plates containing the

166 ... Liu & Zheng

antibiotic streptomycin. The isolation plates were cultivated in the basement (5-10°C,
dark) and at room temperature (about 20°C, natural solar day/night cycle). When
colonies were visible under stereomicroscopes, single colonies were transferred to PDA
slant tubes for subsequent purification and identification.

Cultures

Vouchers of the two strains studied are preserved in culture collections and the
herbarium of the Institute of Microbiology, Chinese Academy of Sciences, Beijing,
China: living cultures in the Culture Collection of the State Key Laboratory of Mycology
(Am-) and the China General Microbiological Culture Collection Center (CGMCC);
and dried cultures in the Herbarium Mycologicum Academiae Sinicae (HMAS).

Media & cultivation

PDA (pH 7) and modified SMA (dextrose 20 g, asparagine 2 g, KH,PO, 0.5 g,
MgSO,°7H,O 0.25 g, thiamine chloride 0.5 mg, agar 20 g in 1000 mL distilled water,
pH 7; Hesseltine & Ellis 1973) were used for morphological studies. PDA was also used
for the mating experiments (with 3% lecithin added) and the detection of maximum
growth temperatures. Cultivation durations and temperatures were: 1) 5-7(-10) days
at 18°C for describing morphological features; 2) 4-7 days at 25-35°C for determining
maximum growth temperatures; and 3) 7-20 days at 18°C for observing sexual reactions.
Seven-day cultures in test tubes were used to measure the height of colonies.

Taxonomy

Ambomucor ovalisporus X.Y. Liu & R.Y. Zheng, sp. nov. FIG. 1
MycoBank MB518427

Differs from all other Ambomucor spp. by its higher colonies, its wider hyphae, its fewer
sporangiophore types, its pyriform to obovoid apophysate fertile sporangia, and its
globose to oval non-vacuolate sporangiospores.

Type: China, Tibet, Nyingchi, Bomé, on wild animal dung, 17 July 2004, Xiao-yong Liu
(Holotype, HMAS 98956-1; isotypes, HMAS 98956-2,-3,-4,-5,-6,-7,-8; ex-type cultures,
Am-14, CGMCC 3.06806).

EtryMo_oey: ovalisporus, referring to the oval shape of the sporangiospores.

CoLonigs on SMA and PDA at first granulate and then floccose, 9 cm diameter
in 7 days at 18°C, up to 2.5 cm high, at first dirty white and then ‘Pinkish
Buff’ (Ridgway XXIX), reverse at first dirty white and then ‘Pinkish Cinnamon’
(Ridgway XXIX). HyPHAE 21.5-66.5(-80.5) um diameter, remaining equal
or narrowed at septa, hyaline to yellow-brown. STOLONs absent. RHIZOIDS
scanty, arising from various parts of the sporangiophores and from fertile and
aborted sporangia. SPORANGIOPHORES erect or recumbent, arising directly
from substrate or hyphae, forming both fertile and aborted sporangia always
in a sympodial pattern, (37-)322-1416(-3476) um long, 5.5-79 um diameter,
usually equal or subequal in width throughout, always constricted just beneath
the terminal fertile sporangia, hyaline to brown, with or without granular
contents, always verrucate; main axes mostly simple, single, rarely with 2-3

Ambomucor sp. & var. nov. (China) ... 167

FiGurE 1. Ambomucor ovalisporus (CGMCC 3.6806). a, Sporangiophore apex triple branches
with an aborted sporangium at the point of separation; b, Single sporangiophore with a fertile
top sporangium (in black); c-d, Rhizoids; e-f, Aborted sporangia showing different sizes;
g-i, Columellae with distinct collars; j, Sporangiospores. Scale bars: a-f = 100 um; g-i = 25 um;
j= 10um.

verticillate branches. SeEpTA appearing frequently in the hyphae and the
main axes of sporangiophores, sporadically in the sporangiophore branches.
ABORTED SPORANGIA proliferating repeatedly on the sides of sporangiophores,
up to 30 or more, transferring their contents forward and forming a short
or long chain of aborted sporangia and terminating in a normal fertile
sporangium, pyriform to obovoid and 20-67.5(-83) x 16-58.5(-—76) um, or
globose to subglobose and (12.5-)24-85 um diameter, verrucate, subhyaline.
FERTILE SPORANGIA borne terminally on the main axes or the branches of
sporangiophores, apophysate, thin-walled, verrucate, globose to subglobose,

168 ... Liu & Zheng

26-163.5 um diameter, dark brown when mature, breaking in larger ones,
persistent in smaller ones. COLUMELLAE forming in the fertile sporangia
only, well developed, verrucate, with a distinct collar, very regular in shape,
hyaline; larger ones subglobose and 102.5-186.5 um diameter, or oblate and
75.5-92(-179) x 55-73.5(-122) um; smaller ones applanate and 34.5-39.5 x
26.5 um. SPORANGIOSPORES forming in fertile sporangia only, released through
breaking or moving backward to the sporangiophores, smooth, mostly oval and
6-12.5 x 4.5-9 um, sometimes globose or subglobose and (5.5-)6.5-9(-10) um
diameter, hyaline. CHLAMYDOSPORES absent. ZYGOSPORES unknown.

ComMENTs: The maximum growth temperature for Ambomucor ovalisporus
Am-14 was determined as 27°C (results of two tests). Crosses of Ambomucor
ovalisporus with A. clavatus and A. seriatoinflatus var. seriatoinflatus, var.
brevior, and var. longior all failed to produce zygospores.

Ambomucor seriatoinflatus var. longior X.Y. Liu & R.Y. Zheng, var. nov.
MycoBank MB518428 Fic. 2
Differs from other Ambomucor seriatoinflatus varieties by its higher colonies, its

abundant aerial hyphae, its longer narrower sporangiophore branches, and its later
sporulation.

Type: China, Shaanxi Province, Taibai Mountain, Shangbansi Temple, in moss and
humus on rock, 11 October 2002, Xue-wei Wang (Holotype, HMAS240174-1; isotypes,
HMAS 240174-2,-3,-4,-5,-6,-7,-8; ex-type cultures, Am-16, CGMCC 3.014179).

Erymo_oey: longior, referring to the sporangiophore primary branches being longer

than those of Ambomucor seriatoinflatus var. seriatoinflatus.
Cotonizs on SMA and PDA floccose, zonate after 3 days, reaching 9 cm
diameter in 6-7 days at 18°C, about 3-5 mm high, at first white, then grayish
white, at last grayish white on the edge and ‘Dark Purplish Gray’ (Ridgway LHI)
in the middle, reverse ‘Olive-Buff’ (Ridgway XL). HypHaz branched, nonseptate
when young, septate in age, 3-7(-8) um diameter in substrate, abundant in
air and (1-)1.5-5(-8) um diameter. SroLons absent. RHIZOIDs scanty, not
opposite sporangiophores, arising from various parts of the sporangiophores
or from the aborted sporangia. SPORANGIOPHORES erect or recumbent, arising
directly from substrate or aerial hyphae, forming fertile or aborted sporangia, of
three main types: 1) bearing normal fertile sporangia only, 2) aborted sporangia
only, or 3) both simultaneously; main axes of the sporangiophores rarely
simple, usually branched, 2.9-7.5 um diameter from substrate, 2-3(-3.5) um
diameter from aerial hyphae; primary branches 1-3, single, in pairs, more often
monopodially or pseudoverticillately rebranching, sometimes sympodially and
branching no more than three times, (15-)60-543(-1600) um long, 2-8.5 um
diameter; main axes and branches more or less straight, or subcurved, usually
equal or subequal in width throughout, hyaline to pale greenish brown, with

Ambomucor sp. & var. nov. (China) ... 169

FiGuRE 2. Ambomucor seriatoinflatus var. longior (CGMCC 3.14179). a-e, Upper portions of
sporangiophores showing various structures: aborted sporangia (blank) and fertile sporangia
(shadowed); f, Rhizoids; g-h, Chlamydospores; i-q, Columellae with distinct collars;
r, Sporangiospores. Scale bars: a~h = 100 um; i-q = 25 um; r= 10 um.

or without granular contents, becoming vacuolate in age. SEPTA in main axes
and branches of the sporangiophores, usually present at the place of branching,
sometimes without definite position. FERTILE SPORANGIA appearing after three
days, borne terminally on the main axes or branches of the sporangiophores,
globose to slightly depressed globose, (21.5-)24.5-47(-53) um diameter in
larger ones on the sporangiophores from substrate, 7-17 um diameter in smaller
ones on the sporangiophores from aerial hyphae, non-apophysate, breaking or
slowly dissolving, thin-walled, dark brown when mature, pale grayish brown
in broken pieces of wall. ABORTED SPORANGIA borne terminally on the main

170 ... Liu & Zheng

axes and branches of sporangiophores, or repeatedly proliferating terminally or
laterally, forming intercalary sporangiophores between two aborted sporangia,
no more than three times, transferring their contents forward until a fertile and
normal sporangium is formed, non-apophysate or slightly apophysate, ovoid
and 10-24 x 7-19 um, globose and 13-22(-26) um diameter, granulate when
young, vacuolate in age, subhyaline. COLUMELLAE formed in fertile sporangia
only, well developed, very regular in shape, ovoid to ellipsoid-ovoid, depressed-
globose to sub-applanate, 6.5-21 x 5-16.5 um, hyaline, some with grayish to
brownish content, smooth, regularly with a distinct collar. SPORANGIOSPORES
forming in fertile sporangia, ellipsoid to broadly oblong-ellipsoid and 3.5-7 x
2-3.5 um, (1-)2-guttulate, hyaline, grayish in mass, smooth, becoming vacuolate
in old cultures. CHLAMYDOSPORES rare, single, or in short chains, subglobose,
ovoid or irregular, 5-18.5 x 3-14 um, hyaline to yellow. ZyGosporEs unknown.

ComMENTs: The maximum growth temperature for Ambomucor seriatoinflatus
var. longior Am-16 was determined as 30°C (results of two tests). Crosses of
Ambomucor seriatoinflatus var. longior with A. clavatus, A. ovalisporus, and
A. seriatoinflatus var. seriatoinflatus and var. brevior all failed to produce
zygospores.

Key to the taxa of Ambomucor
(revised from Zheng & Liu 2014)

1. Colonies up to 25 mm high; hyphae 21.5-66.5 um diameter; both fertile and aborted
sporangia borne on the same sporangiophore; fertile sporangia apophysate;
sporangiospores globose to oval, avacuolate; maximum growth temperature
Dp A stag shy bo tueee cbse shag Er Sabah ht hal bh Ptah obs Fb hy Sea he ral ob hy Pom okt Aa A. ovalisporus

1. Colonies 1-5 mm high; hyphae 1.5-14 «um diameter; fertile or aborted sporangia
or both borne on the same sporangiophore; fertile sporangia non-apophysate;
sporangiospores ellipsoid to oblong-ellipsoid, vacuolate; maximum growth
temperature no less than B0°C ens 5 te ss Ue ae stews a hw saws OE a ee oe Z

2. Aborted sporangia oval to ellipsoidal; columellae of the fertile sporangia mostly
applanate, rarely globose to subglobose; aborted outgrowths frequent, often
spirally twisted; maximum growth temperature 33°C ............. A. clavatus

2. Aborted sporangia globose, subglobose to ovoid; columellae of the fertile sporangia
usually not as above; aborted outgrowths absent or rare, not spirally twisted
when present; maximum growth temperature 30°C ...................0008 3

3. Aborted sporangia borne in chains of up to 20; aborted outgrowths usually absent
and not in a zigzag manner; main axes and branches more or less straight, rarely
SHOE VACUEVED 1 tp. Bess Nak vaso kmh skies 2 A. seriatoinflatus var. seriatoinflatus

3. Aborted sporangia usually not observed exceeding chains of 6; aborted outgrowths
when present formed sympodially; main axes and branches subcurved and
fately: straieht ta: Sub-Strar int! Far 55: Face nas: Bape Pape oP aps eB Pees Pee BSc le Bate 4

Ambomucor sp. & var. nov. (China) ... 171

4. Colonies 3-5 mm high; aerial hyphae abundant; fertile sporangia appearing after
three days; aborted sporangia globose when diameter is less than 26 um;
columellae less than 21 um in width ............ A. seriatoinflatus var. longior

4. Colonies 1-3 mm high; aerial hyphae scarce; fertile sporangia appearing after one
day; aborted sporangia globose, up to 66 um in diameter; columellae 42 um or
TOTS WIGS Pe te coal ei cu Bea teva Mh eva Men td tn At own A A. seriatoinflatus var. brevior

Discussion

Based on the results of studies on several genera of Mucorales, such as
Actinomucor (Zheng & Liu 2005), Cunninghamella (Zheng & Chen 2001),
Pilaira (Zheng & Liu 2009), Rhizomucor (Zheng et al. 2009), and Rhizopus
(Zheng et al. 2007), the range of maximum growth temperatures (MGT) is an
important criterion for helping circumscribe genera and distinguish species;
however, it is not a decisive characteristic and sometimes there is overlap
among taxa. The MGTs of the three Ambomucor species (A. ovalisporus 27°C;
A. seriatoinflatus 30°C; A. clavatus 33°C) occupy a narrow range that supports
their generic afhnity, but with significant differences that support the species
delimitation.

Acknowledgments

The study was partially supported by the National Natural Science Foundation of
China (Nos. 31070019 and 31370068), the Ministry of Science and Technology of China
(No. 2012FY111600) and the Foundation of the Knowledge Innovation Program of the
Chinese Academy of Sciences (No. KSCX2-EW-J-6). We greatly appreciate Associate Prof.
Xue-wei Wang of this department for collecting and isolating the type strain of Ambomucor
seriatoinflatus var. longior. We also thank Ms. Hong-mei Liu of this research group for
preparing the media and plates, helping with isolation, doing mating experiments,
and testing the maximum growth temperatures. Finally we thank Dr. Gerald L. Benny
(University of Florida, Gainesville, U.S.A.) and Dr. Bo Huang (Anhui Agricultural
University, Hefei, P.R. China) for critical comments and presubmission review.

Literature cited

Hesseltine CW, Ellis JJ. 1973. Mucorales. 187-217, in: GC Ainsworth, et al. (eds). The fungi, vol. 4b.
Academic Press, New York.

Zheng RY, Chen GQ. 2001. A monograph of Cunninghamella. Mycotaxon 80: 1-75.

Zheng RY, Liu XY. 2005. Actinomucor elegans var. meitauzae, the correct name for A. taiwanensis
and Mucor meitauzae (Mucorales, Zygomycota). Nova Hedwigia 80: 419-431.
http://dx.doi.org/10.1127/0029-5035/2005/0080-0419

Zheng RY, Liu XY. 2009. Taxa of Pilaira (Mucorales, Zygomycota) from China. Nova Hedwigia 88:
255-267. http://dx.doi.org/10.1127/0029-5035/2009/0088-0255

Zheng RY, Liu XY. 2014 [“2013”]. Ambomucor gen. & spp. nov. from China. Mycotaxon 126:
97-108. http://dx.doi.org/10.5248/126.97

Zheng RY, Chen GQ, Huang H, Liu XY. 2007. A monograph of Rhizopus. Sydowia 59: 273-372.

Zheng RY, Liu XY, Li RY. 2009. More Rhizomucor causing human mucormycosis from China:
R. chlamydosporus sp. nov. Sydowia 61: 135-147.

MY COTAXON

http://dx.doi.org/10.5248/130.173
Volume 130, pp. 173-179 January-March 2015

Perenniporia koreana, anew wood-rotting basidiomycete
from South Korea

YEONGSEON JANG’, SEOKYOON JANG’, YOUNG WOON LIM’,
CHANGMU KIM? & JAE-JIN K1m’*

‘Division of Environmental Science & Ecological Engineering,
College of Life Sciences & Biotechnology, Korea University,
5-1 Anam-dong, Seongbuk-gu, Seoul, 136-701, Korea
School of Biological Sciences, Seoul National University, Seoul, 151-747, Korea
°National Institute of Biological Resources, Environmental Research Complex,
Incheon, 404-708, Korea

*CORRESPONDENCE TO: jae-jinkim@korea.ac.kr

ABSTRACT — A new Perenniporia species, P. koreana, is described based on morphological
and molecular data from specimens collected from Gyeonggi-do and Gangwon-do, Korea.
It is characterized by annual resupinate basidiocarps that are very pale brown to yellow
when dry. Microscopically, it has a dimitic hyphal system with dextrinoid and cyanophilous
skeletal hyphae, and its basidiospores are ellipsoid, hyaline, thick-walled, smooth, dextrinoid,
cyanophilous, (5.9-)6-7(-7.5) x (3.7-)3.9-5.2(- 5.7) um.

Key worps —phylogeny, Polyporaceae, polypore, taxonomy

Introduction

Perenniporia Murrill is a polypore genus of wood-rotting fungi. The
characteristics of this genus are: (1) annual to perennial, resupinate to pileate
basidiocarps; (2) dimitic to trimitic hyphal system with clamped generative
hyphae and non-dextrinoid to strongly dextrinoid or amyloid vegetative
hyphae; and (3) usually thick-walled, ellipsoid to truncate, variably dextrinoid
or amyloid basidiospores (Gilbertson & Ryvarden 1987; Zhao & Cui 2013a,b).
There are about 90 Perenniporia species reported in the world and this genus
was shown to be polyphyletic when evolutionary relationships were inferred
from the representative Perenniporia species including the type, P. medulla-
panis (Jacq.) Donk (Robledo et al. 2009; Zhao et al. 2013; Zhao & Cui 2013a,b).

174 ... Jang & al.

In Korea, nine Perenniporia species have been recorded: P. fraxinea (Bull.)
Ryvarden, P. fraxinophila (Peck) Ryvarden, P. medulla-panis, P. minutissima
(Yasuda) T. Hatt. & Ryvarden, P. ochroleuca (Berk.) Ryvarden, P. ohiensis (Berk.)
Ryvarden, P. subacida (Peck) Donk, P. tephropora (Mont.) Ryvarden, and P
truncatospora (Lloyd) Ryvarden (Lee & Jung 2005; Park & Lee 2011). During
research on indigenous fungi in South Korea, we found eight specimens that
represent Perenniporia but had characteristics that deviated from the previously
reported species. Based on morphological and molecular analyses, we describe
them as a new species.

Materials & methods

Collection and morphological examination

Eight Perenniporia specimens collected from Gyeonggi-do and Gangwon-do
in Korea were used in this study. We examined them macro- and microscopically
according to Jang et al. (2013). Spores were measured in Melzer’s reagent and 5% of
the measurements from both ends are presented in parentheses (Zhao & Cui 2013a).
Color codes follow Munsell (2009). The studied specimens have been deposited at the
Herbarium of National Institute of Biological Resources, Korea (KB).

Phylogenetic analysis

Genomic DNA was extracted from the specimens according to Jang et al. (2012).
PCR reactions were primed with ITSIF (Gardes & Bruns 1993) / ITS4 (White et al.
1990) for internal transcribed spacer (ITS) region and with LROR/LR7 (Vilgalys &
Hester 1990) for nuclear large subunit ribosomal DNA (nLSU) region according to Jang
et al. (2012). DNA was sequenced by Macrogen DNA sequencing service (Seoul, Korea),
and the sequences were deposited in GenBank, NCBI. The ITS and nLSU sequences of
closely related species were retrieved from GenBank. The sequences were aligned with
MAFFT 7.130 (Katoh & Standley 2013) using the L-INS-i method for each dataset. For
Bayesian analysis, the best-fit model for each dataset was calculated by MrModeltest 2.3
(Nylander 2004) under AIC criterion. The two datasets were combined and the selected
models were applied. The 50% majority rule consensus tree was calculated by MrBayes
3.2 (Ronquist et al. 2012) according to Jang et al. (2013).

Taxonomy

Perenniporia koreana Y. Jang & J.J. Kim, sp. nov. Fic. 1
MycoBank MB 807626

Differs from other Perenniporia species by its annual resupinate basidiocarps with grayish
orange pore surface, lack of rhizomorphs in most specimens, a dimitic hyphal system
with dextrinoid skeletal hyphae, and ellipsoid non-truncate dextrinoid basidiospores.

Fic. 1. Perenniporia koreana (KB NIBRFG107080, holotype). A. Basidiocarp. Scale bar = 1 cm.
B. Microscopic features: a, basidioles and basidia; b, cystidioles; c, basidiospores; d, generative
hyphae from trama; e, generative hyphae from context; f, skeletal hyphae from trama; g, skeletal
hyphae from context.

koreana sp. nov. (South Korea) ... 175

iporia

Perenn

176 ... Jang & al.

Type: Korea. Kyeonggi-do, Dongducheon-si, Mt. Soyo, 37°56’44”N 127°04’08’E, on
the branch of Quercus sp., 17 May 2008, Jae-Jin Kim KUC20080517-15 (Holotype, KB
NIBRFG107080; GenBank KJ156301, KJ156309).

Erymo.oey: The locality (Korea) of the type specimen.

BASIDIOCARPS annual, resupinate, adnate, ca. 8.5 cm or more in longest
dimension, ca. 4 cm in widest dimension. Sterile margin felty, orange gray
(10YR8/2), up to 1 cm wide, usually without rhizomorphs (present only
in KB NIBRFG107396). Pores round to angular, 5-6 per mm; dissepiments
thin, entire; pore surface grayish orange (1OYR8/3-8/4) to brownish orange
(10YR7/6-7/8) in dried condition. Context corky, cream to buff, up to 0.5 mm
thick. Tubes corky, concolorous with pore surface, up to 2 mm thick.

HyPHAL SYSTEM dimitic; generative hyphae with clamp connections; skeletal
hyphae dextrinoid, cyanophilous.

CONTEXT generative hyphae hyaline, thin-walled, 1.5-3 um in diameter;
skeletal hyphae dominant, hyaline, thick-walled, with a distinct lumen,
interwoven, 3.5-6 um in diameter.

TUBES generative hyphae hyaline, thin-walled, 1.5-2.5 um in diameter;
skeletal hyphae dominant, hyaline, thick-walled, with a narrow lumen to
subsolid, rarely branched, interwoven, 1.5-4.5 um in diameter. Cystidia
absent, fusoid cystidioles present, 19.5-23 x 5-6.5 um; basidia barrel-shaped,
4 sterigmate, 21-23 x 7.5-9 um.

Basiprospores ellipsoid, hyaline, thick-walled, smooth, dextrinoid,
cyanophilous, (5.9-)6-7(-7.5) x (3.7-)3.9-5.2(-5.7) um, L = 6.56, W = 4.45
(n = 120/4).

ECOLOGY & DISTRIBUTION — Korea, on hardwoods causing white rot.

ADDITIONAL SPECIMENS EXAMINED: KOREA, KYEONGGI-DO, Uijeongbu-si, Mt.
Dobong, on fallen wood branch, 30 October, 2009, Yeongseon Jang KUC20091030-
32 (KB NIBRFG113564; GenBank KJ156305, KJ156313); Yongin-si, Mt. Gwanggyo,
37°21'05"'N 127°03’14’E, on hardwood, 14 August 2012, Yeongseon Jang KUC20120814-
17 (KB NIBRFG125415; GenBank KJ156306, KJ156314); Yangju-si, Mt. Goryeong, on
wood, 11 July 2009, Yeongseon Jang KUC20090711-70 (KB NIBRFG113613; GenBank
KJ156304, KJ156312); Dongducheon-si, Mt. Soyo, 37°56’44”N 127°04’08’E, on the
branch of hardwood, 17 May 2008, Jae-Jin Kim KUC20080517-02 (KB NIBRFG107071;
GenBank KJ156300, KJ156308); GANGWON-DO, Heongseong-gun, Mt. Chiak, on wood,
4 April 2009, Young Woon Lim KUC20090404-27 (KB NIBRFG113243; GenBank
KJ156303, KJ156311); 37°20’16”N 128°03’27”E, on hardwood, 2 October 2008, Jae-Jin
Kim KUC20081002J-02 (KB NIBRFG107396; GenBank KJ156302, KJ156310); Wonju-
si, Mt Chiak, 37°21’43”N 128°02’36’E, on the branch of Quercus sp., 10 May 2008, Jae-
Jin Kim KUC20080510-01 (KB NIBRFG107760; GenBank KJ156299, KJ156307).

Phylogeny
The combined ITS+nLSU dataset contained 56 taxa and 2016 characters, of
which the ITS dataset was 667 characters and nLSU was 1349 characters. By the

Perenniporia koreana sp. nov. (South Korea) ... 177

1 Perenniporiella chaquenia MUCL 47648 (FJ411084/FJ393856)
1 Perenniporiella pendula MUCL 46034 (FJ411081/FJ393853)
258 Perenniporiella micropora MUCL 43581 (FJ411086/FJ393858)
0.96 Perenniporia minor Cui 5782 (HQ883475/HQ6541 15)
1p Perenniporia lacerata Cui 7220 (JX141448/JX141458)
Perenniporia macropora Zhou 407 (JQ861746/JQ861 762)
968 Perenniporia tibetica Cui 9457 (JF706326/JF706332)
Perenniporia nanlingensis Cui 7620 (HQ848477/HQ848486)
0.97 Perenniporia japonica Cui 7047 (HQ654097/HQ6541 11)
0.79 Perenniporia ochroleuca Dai 11486 (HQ654105/JF706349)
0.82 4 Perenniporia detrita MUCL 42649 (FJ411099/FJ393866)
0.94 Perenniporia ohiensis Cui 5714 (HQ654103/HQ654116)
1 Abundisporus sclerosetosus MUCL 41438 (FJ411101/FJ393868)
Abundisporus violaceus MUCL 38617 (FJ411100/FJ393867) |
1 Perenniporia substraminea Cui 10177 (JQ001852/JQ001 844)

0.66 4 Perenniporia medulla-panis MUCL 43520 (FJ411087/FJ393875)
O87 Perenniporia hainaniana Cui 6364 (JQ861743/JQ861759)
Perenniporia straminea Cui 8718 (HQ876600/JF 706335)
222 093 Perenniporia piceicola Dai 4184 (JF706328/JF 706336)

Perenniporia russeimarginata Yuan 1225 (JQ861749/JQ861 765)
1p Perenniporia truncatospora Cui 6987 (JN048778/HQ6541 12)
Perenniporia pyricola Dai 10265 (JN048761/JN048781)
0.51 9 Perenniporia tephropora Cui 6331 (HQ848473/HQ848484)
Perenniporia subtephropora Dai 10962 (JQ861752/JQ861 768)
0.99 Perenniporia aridula Dai 12396 (JQ001854/JQ001846)
Perenniporia tenuis Wei 2783 (JQ001858/JQ001848)
4 Perenniporia maackiae Cui 5605 (JN048760/JN048780)
Perenniporia corticola Dai 7330 (HQ654094/HQ654108)
Perenniporia koreana KUC20090711-70 (KJ156312, KJ156304)
°97) Perenniporia koreana KUC20080517-15 (KJ156309, KJ156301) - holotype
Perenniporia koreana KUC20080517-02 (KJ156308, KJ156300)
ip Perenniporia koreana KUC20090404-27 (KJ156311, KJ156303)
1 1] | Perenniporia koreana KUC20081002J-02 (KJ156310, KJ156302)
Perenniporia koreana KUC20120814-17 (KJ156314, KJ156306)
Perenniporia koreana KUC20091030-32 (KJ156313, KJ156305)
4 Perenniporia koreana KUC20080510-01 (KJ156307, KJ156299)
1p Perenniporia luteola Harkonen 1308b (JX141457/JX141467)
; 0.99 Perenniporia luteola Harkonen 1308a (JX141456/JX141466)
1 Perenniporia rhizomorpha Dai 7248 (JF706330/JF706348)
Perenniporia rhizomorpha Cui 7507 (HQ654107/HQ654117)
1p Perenniporia bannaensis Cui 8562 (JQ291728/JQ291 730) | |
Perenniporia bannaensis Cui 8560 (JQ291727/JQ291729)
yeh 4 Perenniporia subacida MUCL 31402 (FJ411103/FJ393880)
Perenniporia narymica Dai 10510 (HQ654101/JF 706346)
1 Perenniporia subadusta Cui 8459 (HQ876606/HQ654113)
O74 Microporellus violaceo-cinerascens MUCL 45229 (FJ411106/FJ393874)
oes Perenniporia martia MUCL 41677 (FJ411092/FJ393859)
1--Perenniporia fraxinea Cui 8871 (JF706329/JF706345)
i Perenniporia robiniophila Cui 9174 (HQ876610/JF706343)
4 Perenniporia vicina MUCL 44779 (FJ411095/FJ393862)
7 Perenniporia formosana Dai 5245 (HQ876612/JX941590)
051 Perenniporia tianmuensis Cui 2648 (JX141453/JX141463)
1 Perenniporia contraria Knudsen 04-111 (JQ861737/JQ861755)
Perenniporia fergusii Gilbertson 16116 (HQ876607/JF 706337)
1 Pyrofomes demidoffii MUCL 41034 (FJ411105/FJ393873)
Donkioporia expansa MUCL 35116 (FJ411104/FJ393872)

0.01

Fic. 2. 50% majority consensus tree of Perenniporia koreana and allied species inferred from
combined ITS+nLSU region sequences. Posterior probabilities over 0.5 are shown above branches.
GenBank accession numbers are presented in parentheses (ITS/nLSU). Perenniporia koreana is
indicated by bold type.

model selection, GTR+I+G model was chosen for both ITS and nLSU datasets.
Pyrofomes demidoffii (Lév.) Kotl. & Pouzar (MUCL 41034) and Donkioporia
expansa (Desm.) Kotl. & Pouzar (MUCL 35116) were used as outgroup (Zhao
& Cui 2013a,b; Zhao et al. 2013). The phylogenetic tree (Fic. 2) provides
the sequence information for the included taxa. Our analysis supported two
main groups, I and II. Group I, which contained Perenniporia species with
typically truncate basidiospores, included P. medulla-panis (the type species)
as well as Abundisporus sclerosetosus Decock & Laurence, A. violaceus (Wakef.)
Ryvarden, Perenniporiella chaquenia Robledo & Decock, P pendula Decock &
Ryvarden, and P. micropora (Ryvarden) Decock & Ryvarden. Group IJ included

178 ... Jang & al.

Perenniporia species with non-truncate basidiospores (except for P fergusii
Gilb. & Ryvarden with ellipsoid to slightly truncate basidiospores; Gilbertson
& Ryvarden 1987), and Microporellus violaceocinerascens (Petch) A. David &
Rajchenb. Perenniporia koreana was in group II and monophyletic with high
posterior probability value (1.0 p.p.). Its sister taxa were P. luteola B.K. Cui &
C.L. Zhao and P. rhizomorpha B.K. Cui et al., and these three taxa clustered
with P bannaensis B.K. Cui & C.L. Zhao, P. subacida, and P. narymica (Pilat)
Pouzar in a clade with high support (1.0 p.p.).

Discussion

Morphologically, Perenniporia koreana is very similar to its sister taxa,
P. luteola and P. rhizomorpha, which are also characterized by a resupinate
basidiocarp, yellowish pore surface, dextrinoid skeletal hyphae, and dextrinoid
non-truncate basidiospores. However, P. luteola differs by its perennial habit
and slightly wider basidiospores (6.1-6.9 x 5.1-5.4 um; Zhao & Cui 2013a),
and P. rhizomorpha differs by having typical rhizomorphs, slightly longer
basidiospores (5.3-6.5 x 4.1-5.2 um), contextual skeletal hyphae encrusted
with fine crystals and lacking cystidia and cystidioles (Cui et al. 2007).

Perenniporia tenuis (Schwein.) Ryvarden and P. bannaensis are similar to
P. koreana in their annual resupinate basidiocarps, yellowish pore surface,
dimitic hyphal system with dextrinoid skeletal hyphae, fusoid cystidioles, and
dextrinoid basidiospores. However, P. tenuis differs in its larger pores (3-5/mm)
and small ellipsoid to truncate basidiospores (4.5-6 x 3.5-4.5 um; Ryvarden
& Gilbertson 1994), while P bannaensis differs in its small pores (6-8/mm),
entire to lacerate dissepiments, and shorter basidiospores (5.2-6 x 4-4.6 um;
Zhao et al. 2013).

Phylogenetically (Fic. 2), the species relationships in our analysis differ
substantially from those of Zhao et al. (2013) and Zhao & Cui (2013a,b). Our
analysis clustered Perenniporia luteola, PR. rhizomorpha, and P. bannaensis in
group II, a relationship also observed when P. koreana was excluded from
the phylogenetic analysis (not shown). In contrast, P rhizomorpha and
P. bannaensis were monophyletic with P. medulla-panis in Zhao et al. (2013);
and P. luteola, P. rhizomorpha, and P. bannaensis clustered with P medulla-panis
in Zhao & Cui (2013a,b). Further research with additional loci would help to
resolve the species relationships of Perenniporia.

In conclusion, we propose Perenniporia koreana as a new species with only
minute morphological distinction but with distinct genetic differences.

Acknowledgments

This research was supported by Basic Science Research Program through the
National Research Foundation of Korea (NRF) funded by the Ministry of Education
(NRF-2013R1A1A2A10011390) and was supported by the Indigenous Species Survey

Perenniporia koreana sp. nov. (South Korea) ... 179

and Investigation project from the National Institute of Biological Resources (NIBR)
under the Ministry of Environment, Republic of Korea. We are much obliged to Dr.
Bao-Kai Cui and Dr. Tsutomu Hattori for their valuable suggestions on the manuscript.

Literature cited

Cui BK, Dai YC, Decock C. 2007. A new species of Perenniporia (Basidiomycota, Aphyllophorales)
from eastern China. Mycotaxon 99: 175-180.

Gardes M, Bruns TD. 1993. ITS primers with enhanced specificity for basidiomycetes — application
to the identification of mycorrhizae and rusts. Mol. Ecol. 2: 113-118.
http:/dx.doi.org/10.1111/j.1365-294X.1993.tb00005.x

Gilbertson RL, Ryvarden L. 1987. North American polypores, vol. 2. Fungiflora, Oslo.

Jang Y, Choi HE, Lim Y W, Lee JS, Kim J-J.2012. The first report of Ceriporia lacerata (Phanerochaetaceae,
Basidiomycota) in Korea. Mycotaxon 119: 397-403. http://dx.doi-org/10.5248/119.397

Jang Y, Lee SW, Lim YW, Lee JS, Hallenberg N, Kim J-J. 2013. Hypochnicium pini, a new corticioid
basidiomycete in East Asia. Mycotaxon 124: 209-217. http://dx.doi.org/10.5248/124.209

Katoh K, Standley DM. 2013. MAFFT Multiple sequence Alignment Software Version
7: Improvements in Performance and Usability. Mol. Biol. Evol. 30: 772-780.
http://dx.doi.org/10.1093/molbev/mst010

Lee JS, Jung HS. 2005. List of recorded Korean Aphyllophorales. Kor. J. Mycol. 33: 38-53.

Munsell color. 2009. Munsell soil color charts with genuine Munsell color chips. Grand Rapids,
Michigan, U.S.A

Nylander JAA. 2004. MrModeltest v2. Evolutionary Biology Center, Uppsala University, Uppsala,
Sweden.

Park W, Lee J. 2011. New wild Fungi of Korea. Kyohak Publishing, Korea.

Robledo GL, Amalfi M, Castillo G, Rajchenberg M, Decock C. 2009. Perenniporiella chaquenia
sp. nov. and further notes on Perenniporiella and its relationships with Perenniporia (Poriales,
Basidiomycota). Mycologia 101: 657-673. http://dx.doi.org/10.3852/08-040

Ronquist F, Teslenko M, Mark P van der, Ayres D, Darling A, Hohna S, Larget B, Liu L, Suchard MA,
Huelsenbeck JP. 2012. MrBayes 3.2: Efficient Bayesian phylogenetic inference and model choice
across a large model space. Syst. Biol. 61: 539-542. http://dx.doi.org/10.1093/sysbio/sys029

Ryvarden L, Gilbertson RL. 1994. European polypores 2. Synopsis fungorum 7. Fungiflora, Oslo.

Vilgalys R, Hester M. 1990. Rapid genetic identification and mapping of enzymatically amplified
ribosomal DNA from several Cryptococcus species. J. Bacteriol. 172: 4238-4246.

White TJ, Bruns TD, Lee S, Taylor JW. 1990. Amplification and direct sequencing of fungal
ribosomal RNA genes for phylogenetics. 315-322, in: MA Innis et al. (eds). PCR Protocols: a
Guide to Methods and Applications. Academic Press, New York.

Zhao CL, Cui BK. 2013a. Three new Perenniporia (Polyporales, Basidiomycota) species
from China based on morphological and molecular data. Mycoscience 54: 231-240.
http://dx.doi.org/10.1016/j.myc.2012.09.013

Zhao CL, Cui BK. 2013b. Morphological and molecular identification of four new resupinate
species of Perenniporia (Polyporales) from southern China. Mycologia 105: 945-958.
http://dx.doi.org/10.3852/12-201

Zhao CL, Cui BK, Dai YC. 2013. New species and phylogeny of Perenniporia based on morphological
and molecular characters. Fungal Divers. 58: 47-60. http://dx.doi.org/10.1007/s13225-012-0177-6

MY COTAXON

http://dx.doi.org/10.5248/130.181
Volume 130, pp. 181-190 January-March 2015

New records of corticolous myxomycetes from Turkey

R. BATUR ORAN! & C. CEM ERGUL"

Department of Biology, Faculty of Arts & Sciences, Uludag University,
16059, Bursa, Turkey

*CORRESPONDENCE TO: ergulc@uludag.edu.tr

ABSTRACT- Eight myxomycete species from the Marmara region are newly recorded for
Turkey: Amaurochaete tubulina, Comatricha longipila, Diderma cinereum, D. umbilicatum,
Didymium lenticulare, Physarum serpula, Stemonitis inconspicua, and Symphytocarpus
confluens.

Keyworps- Quercus bark, moist chamber technique

Introduction

The Marmara region of Turkey is located in the Euro-Siberian and the
Mediterranean phytogeographical regions of Turkey. The western part of the
region is subjected to Mediterranean climate and is covered with pine forests
and shrubs, whereas the eastern and northern parts have a continental climate
and are rich in oak forests (Atalay 2002).

Turkey is an important world distribution area for Quercus diversity (Gtinal
1997). There are 18 species of oak trees in Turkey (Segmen et al. 1995), of which
11 occur in the Marmara region.

The first myxomycetes for Turkey were reported by Harkénen & Uotila
(1983) and Harkonen (1987). Although the number of myxomycete species
worldwide is about 1000 (Lado 2001), the reported numbers for Turkey
are relatively low, around 232 (Sesli & Denchev 2014). With this paper we
contribute further knowledge on the myxomycetes in the country by presenting
eight species collected on bark of Quercus trees in the Marmara region, all new
records for Turkey.

Materials & methods
Field excursions were carried out in a randomised manner during June and October
of 2005-2008. All material was collected from the bark of Quercus species in different

182 ... Oran & Ergiil

provinces of Turkey’s Marmara region. Myxomycete fructifications were obtained from
moist chamber cultures in the laboratory as described in Stephenson & Stempen (1994).
The cultures were moistened with distilled water and kept under diffuse sunlight within
a temperature range of 22-25°C.

The cultures were examined under a dissecting microscope every day for a 4-week
period. Whenever developing fructifications were found, the bark pieces on which they
were found were removed from the Petri dish, allowed to dry slowly, and then placed
in herbarium boxes. The same chambers were then rewetted for another 4-week period
and examined as before. The specimens are stored in the authors’ personal collections
in the Department of Biology, Uludag University, Bursa, Turkey. Species have been
identified according to Castillo et al. (1997), Farr (1976, 1981), Ing (1999), Keller &
Brooks (1977), Lado & Pando (1997), Martin & Alexopoulos (1969), Mitchell (2013),
Nannenga-Bremekamp (1991), Stephenson & Stempen (1994), and Thind (1997).
Nomenclature follows Lado (2001).

Species recorded

Amaurochaete tubulina (Alb. & Schwein) T. Macbr. PLATE 1
Sporocarps aethaliate. Aethalia pulvinate, somewhat depressed. Sporothecae

covered by a thin, translucent cortex. Hypothallus thin, shining, prominent.
Capillitium branched, irregularly sprouting out from the base, branching and
anastomosing freely. Spore mass black. Spores pale olivaceous, 12-15 um
diam., finely warted.

SPECIMEN EXAMINED - ‘TURKEY, BILECIK/MERKEZ; Karadere-Sitliik road,

40°05’35.3”N 29°49’51.9”E, alt. 977 m, on bark of Quercus frainetto in oak forest,

22.7.2007, Oran 303-1.
COMMENTS—Amaurochaete is a typical genus in the order Stemonitales
characterized by a dendroid capillitium. Previously only a single species,
A. atra (Alb. & Schwein.) Rostaf., has been reported from Turkey (Sesli &
Denchev 2014). An examination of published records of A. tubulina indicates
that this species is frequently isolated from conifer bark, whereas our specimen
was recovered from Quercus bark in a moist chamber culture. Our specimen
appears to represent the first aethaliate structure of an Amaurochaete obtained
from a moist chamber culture.

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PiaTE 1. Amaurochaete tubulina. I- aethalium; II- a. capillitium, b. spores; III- spores.

Corticolous myxomycetes new for Turkey ... 183

Amaurochaete tubulina somewhat resembles A. atra and A. comata G. Lister
& Brandza. It differs from A. atra, which is clearly distinguished by an early
fugacious aethelial wall, in its capillitial structure that widens in the axils and
is connected to the columella but not connected to neighbouring sporangia.
Amaurochaete tubulina is distinguished from A. comata by its sparsely
anastomosing free ends, spiny capillitia, and spores that are less strongly warted
on one side.

PLATE 2. Comatricha lenpipita:| I- sporocarps; I- a. are columella; III- a. columella; b.
capillitium; c. spinose capillitial ends; [V- a. columella, b. capillitium, c. spore; d. capillitial ends;
V- a. capillitium, b. reticulate spores.

Comatricha longipila Nann.-Bremek. PLATE 2

Sporocarps in small groups or solitary, 1.5-2.0 mm tall, nearly erect.
Hypothallus discoid or continuous under the group, brown and inconspicuous.
Stalk up to 33% of the total height, with a net of fibres at the base, often opaque
above. Sporotheca cylindrical or obovoid, 0.3-0.6 mm diam., brown. Peridium
usually completely fugacious but sometimes persistent as a collar around the
stalk apex. Columella merging into the capillitium just below the apex of the
sporotheca. Capillitium primary branches perpendicular to the columella,
hardly anastamosing, dichotomously branched with many, rather straight,
long, free ends at the periphery which are sometimes slightly swollen. Spores
pale red-brown, 6.0-7.0 um diam., finely warted.

SPECIMEN EXAMINED — TURKEY, KIRKLARELI/MERKEZ; Uskiip—Beypinari road 2 km,

41°45’05.7”N 27°26'32.4’E, alt. 403 m, on bark of a solitary Quercus pubescens tree in
agricultural land, 15.6.2006, Oran 91-2.

184 ... Oran & Ergiil

COMMENTS—Our specimen, as was the case for the type specimen, appeared
in a moist chamber culture prepared with a bark sample from Quercus. The
peridium occasionally remains as a permanent collar around the stalk. As
noted by Nannenga-Bremekamp (1991), “C. longipila is characterized by the
lax internal capillitium, with long free ends at the periphery and its small
spores.” However, Ing (1999) indicated that its small spores, lack of an outer
capillitial network, and long free ends separate Comatrica longipila from small
specimens of C. laxa Rostaf.

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PLATE 3. Diderma cinereum. I- individual sporocarps;
I- a. capillitium, b. peridium, c. spores; III- verruculose spores.
Diderma cinereum Morgan PLATE3

Sporocarps sessile, gregarious, subglobose, somewhat depressed, 0.3-0.5
mm diam., pearl-grey. Hypothallus inconspicuous. Peridium single, thin,
smooth, crustose. Capillitium of slender, dark threads, sparsely branched,
easily detached from the columella. Spore-mass black. Spores dark violet-grey,
9-11 um diam., verruculose.

SPECIMEN EXAMINED — TURKEY, BALIKESIR/MERKEZ; Konakpinar-Sépk6y road 5

km, 39°25’00.8”N 27°50’06.5”E, alt. 331 m, on bark of Quercus cerris, 28.7.2006, Oran

242-2.
ComMMENTS— There are various reports of the presence of a single or double
peridium in this species, but our specimen clearly has a single peridium. Ing
(1999) observed that D. cinereum is similar to D. spumarioides (Fr.) Fr. but
differs in having a single peridium, deciduous capillitium, and dark grey spores.

Diderma umbilicatum Pers. PLATE 4
Sporocarps short-stalked or sessile, pearl-grey. Peridium double, the outer
layer calcareous or occasionally limeless, adhering closely to the membranous
inner layer. Dehiscence irregular, leaving a ragged cup which shows white on
the inside. Columella prominent, pale, subglobose, smooth or rugose. Stalk
stout, reddish-brown, covered with white lime. Spores dark brown in mass,
purple brown in transmitted light, densely verruculose, 10-12 um diam.

SPECIMENS EXAMINED — TURKEY, CANAKKALE/YENICE; around Gonen dam,
39°57’41.1”N 27°25'52.2’E, alt. 191 m, on bark of Quercus cerris in composite forest,

Corticolous myxomycetes new for Turkey... 185

PiateE 4. Diderma umbilicatum. I- individual sporocarp;
II- a. peridial fragments, b. capillitium, c. spores; II- a. capillitium, b. spores.

29.7.2006, Oran 253-1; SAKARYA/GEYVE; Bozcagiz-Koru road, 4 km to Koru village,

40°3133”N 30°26’49’E, alt. 350 m, on bark of Quercus virgiliana in oak—-beech forest,

7.10.2007, Oran 266-1.
ComMENtTS—Diderma umbilicatum is included in Diderma subg. Leangium
(Link) Macbr. and has often been considered a variety of D. radiatum
(L.) Morgan (Nannenga-Bremekamp 1991). Ing (1999) suggested that
D. umbilicatum resembles both D. montanum (Meyl.) Meyl. and D. radiatum,
especially with respect to peridial characteristics. Hence, it was considered to
be synonymous with D. radiatum until quite recently. However, it is actually
quite different, having a number of features (e.g., irregular dehiscence, non-
cartilaginous peridium, pale columella, stout stalk) that are not present in
D. radiatum. Under a hand lens, it may resemble D. montanum, from which
it can be distinguished by its thicker and reddish short stalk, large columella,
and usually larger spores. Ing (1999) noted that its large pale sporangia,
conspicuous pale columella, and thick stalk easily separate D. umbilicatum
from other Diderma taxa.

Didymium lenticulare K.S. Thind & T.N. Lakh. PLATE 5

Sporocarps stipitate, up to 2 mm tall, scattered to loosely gregarious.
Hypothallus dark brown, rotate. Stalk 1-1.8 mm long, erect, grooved, tapered,
dark brown below, lighter and translucent above, scantily charged with lime
crystals, particularly at the base. Sporotheca erect or nodding, strongly
discoid, deeply umbilicate at the base, white, 0.5-0.8 mm diam. Peridium
membranous, hyaline, densely covered with lime deposits which form very
irregular ridges and plates. Columella absent. Capillitium of stout, subhyaline
threads, sparingly branched and anastomosed and with bead-like thickenings.
Spore-mass black. Spores deep violaceous brown by transmitted light, globose,
distinctly verrucose, also marked by conspicuous clusters of darker and bigger
warts, 9-11 um diam.

SPECIMEN EXAMINED — TURKEY, KIRKLARELI/PINARHISAR; Cayirdere—Akoren road
2 km, 41°42’19.3’N 27°31'15.7’E, alt. 269 m, on bark of Quercus frainetto in an oak
stand, 25.7.2006, Oran 179-2.

186 ... Oran & Ergiil

PiateE 5. Didymium lenticulare. I- individual sporocarp;
II- a. stipe, b. pseudocolumella, c. capillitial threads; d. spores;
III- a. crusted lime, b. capillitium, c. spores.

ComMMENTS—Didymium lenticulare very closely resembles D. clavus (Alb. &
Schwein.) Rabenh. but differs in a number of features, including its hyaline,
crustose peridium with thick ridges, longer stipe, more dispersed lime crystals
(especially in the lower region), lack of a columella, and spores that are larger,
lighter in color, and distinctly verrucose (Thind 1977).

This species also resembles D. squamulosum (Alb. & Schwein.) Fr. in its
possession of a hyaline peridium and similar spores but differs in its densely
thick calcareous deposits on the peridium, strongly discoid sporotheca, and
lack of a columella. In addition, its stipe is much longer and darker than that
typically observed in D. squamulosum. ‘The stipe of D. lenticulare is non-
calcareous, although it is sprinkled with lime crystals.

The stipe also differs from that of D. iridis (Ditmar) Fr., which has strongly
discoid sporangia that are deeply umbilicate below. The extraordinarily small
lime crystals of D. lenticulare also serve to distinguish it from allied species
(Thind 1977).

eS 3 ¢

‘ge fe. 6

PLatE 6. Physarum serpula. I- plasmodiocarp;
II- a. lime node, b. spores, c. peridium; III- a. lime node, b. spores.

Physarum serpula Morgan PLATE 6

Plasmodiocarpous, forming lines, rings, or a simple network, 0.2-0.4
mm wide, sometimes fusing laterally to form a broad fruiting surface, often
interspersed with globose sporocarps, dull yellow or ochraceous, sessile.
Hypothallus diffuse. Peridium single, thin, of closely encrusted lime globules,

Corticolous myxomycetes new for Turkey ... 187

without calcareous scales, fragile, membranous, persistent. Capillitium dense,
calcareous, the nodes numerous, large, angular, branching, pale yellow or
whitish, connected by short, hyaline threads or broad limy strands and then
almost badhamioid. Spore-mass dull black. Spores dark brown, verruculose,
with a paler and smoother area on one side, 10-13 um diam.

SPECIMEN EXAMINED - TURKEY, Bursa/NILUFER; around Alaaddinbey village,

40°12’10.7”N 28°53’45.2”E, alt. 78 m, on bark of a solitary Quercus robur tree in

agricultural land, 16.9.2007, Oran 312-2.
CoMMENTS—Physarum serpula was treated by Ing (1999) as part of a trio with
P. auriscalpium Cooke and P. decipiens M.A. Curtis, and Martin & Alexopoulos
(1969) observed that these three morphologically similar species were often
confused with one another. Physarum serpula is frequently confused with
P. auriscalpium, which differs by having a peridium dotted with prominent
glossy lime scales rather than the peridium evenly impregnated with lime in
P. serpula. Ing (1999) observed that P decipiens, which is more sporangiate than
P. auriscalpium or P. serpula, has a more badhamioid capillitium.

PLATE 7. Stemonitis inconspicua. 1) sporocarps; II- individual sporocarp; II- apical end of
sporotheca; IV- basal end of sporotheca; V-— inner net of capillitium; VI- surface net.

Stemonitis inconspicua Nann.-Bremek. PLATE 7

Sporocarps clustered on a common hypothallus, sometimes gregarious, total
height 2-8 mm. Hypothallus thin, silvery, red-brown by light microscope. Stalk
approximately 0.5 mm long, thin, black, opaque, shining. Sporothecae brown,
blunt cylindrical, 1.5-2.5 mm tall. Peridium fugacious, except sometimes at
the base, where red-brown fragments show a structure similar to that of the
spores. Columella tapered, dissipating just below the apex. Capillitium forming
approximately 3 meshes on the radius, consisting of slender threads variously

188 ... Oran & Ergiil

widened at the junctions, becoming extremely fine towards the periphery, the
surface net slender, irregular, fragmentary, with spiny free ends, dissipating
towards the apex. Spore mass dark brown. Spores pale red-brown, prominently
banded-reticulate with 2-3 meshes across the hemisphere, 8-9 um diam.
excluding the bands, which form a border approximately 0.5 um high in optical
section.

SPECIMENS EXAMINED — TURKEY, BILEcIK/MERKEZ; Karadere-Sitliik road, 3 km

to Siitltik, 40°05’35.3”N 29°49’51.9’E, alt. 977 m, on bark of Quercus frainetto in an

oak stand, 22.7.2007, Oran 3-1; CANAKKALE/MERKEZ; Can-Canakkale road, Kocalar

turnout, 40°02’03.7”N 26°46’57.1’E, alt. 469 m, on bark of Quercus frainetto in an oak

stand, 6.7.2005, Oran 303-4.
COMMENTS—Stemonitis inconspicua can be distinguished from the similar
S. foliicola Ing by its small size and banded-reticulate spores (Ing 1999).

os a
ae Ow

PLATE 8. Symphytocarpus confluens. I- pseudoaethalium; II- silver coloured hypothallus;
II- a. hypothallus and stipe, b. capillitium; IV- a. enlargements on capillitium, b. spores; V) spores.

Symphytocarpus confluens (Cooke & Ellis) Ing & Nann.-Bremek. PLATE 8

Pseudoaethalium 2-3 mm tall and 0.5-4 cm diam., deep black. Sporothecae
approximately 0.5 mm diam. Hypothallus silvery. Peridium fugacious except
for a number of more or less rounded plates, smooth and connected to the
capillitium. Columella absent or, if present, opaque, dissipating below the apex.
Capillitium a wide-meshed reticulum of thick, dark purple threads with small
axillary membranes, looped at the periphery. Spores dark purple-brown in
transmitted light, (10-)11-13 um diam., distinctly warted.

Corticolous myxomycetes new for Turkey ... 189

SPECIMEN EXAMINED — TURKEY, KIRKLARELI/MERKEZ; Derek6y-Kula road, around

Gegitagzi village, 41°57’34.9"N 27°21'43.4”E, alt. 624 m, on bark of Quercus petraea,

16.6.2006, Oran 110-1.
CoMMENTS—Symphytocarpus confluens, which is close to S. impexus Ing &
Nann.-Bremek, differs in its shorter, more elongated, deep black sporangia,
which occasionally possess an opaque columella that dissipates below the
apex of the sporangium (although it is not uncommon for the columella to be
absent). Hence, the sporangia of S. confluens are much deeper black in color
and have much larger spores that are at least 10 (usually =12) um in diameter
(Nannenga-Bremekamp 1991).

Discussion

Bark from Quercus trees sampled from 11 cities and six provinces in the
Marmara region were the source of our new records for Turkey: Balikesir
province (Diderma cinereum), Bilecik province (Amaurochaete tubulina,
S. inconspicua), Canakkale province (D. umbilicatum, S. inconspicua),
Bursa province, (P. serpula), Kirklareli province (C. longipila, D. lenticulare,
S. confluens), and Sakarya province (D. umbilicatum). As the distribution
shows, two species (Didymium umbilicatum, Stemonitis inconspicua) were
recovered from two different provinces.

The character of the tree bark structure is an important factor for myxomycete
distribution and productivity. Harkonen et al. (2004) concluded that a smooth
basic bark has a low water holding capacity and does not support a high
diversity of myxomycetes, bryophytes, and lichens. McHugh (1998) noted that
individual Quercus species and Quercus forests are the most productive sources
for myxomycetes. Bark from oak trees offers a suitable structure for sheltering
myxomycete spores. Stephenson (1989) noted that the considerable variation in
bark structure offers good potential microhabitats for corticolous myxomycetes.
High water holding capacities may benefit corticolous myxomycetes directly as
their active plasmodia are highly dependent on the availability of liquid water.

Acknowledgements

We thank Prof. S.L. Stephenson (University of Arkansas) and D.W. Mitchell for their
invaluable suggestions to our manuscript. We offer also sincere thanks for the effort and
meticulous attention to the nomenclature provided Dr. S.R. Pennycook. We thank also
AJE for article editing process.

Literature cited

Atalay I. 2002. Ecoregions of Turkey. Meta Basimevi, Izmir, Turkey.

Castillo A, Ilana C, Moreno G. 1997. A critical study of some Stemonitales. Mycological Research
101(11): 1329-1340. http://dx.doi.org/10.1017/S0953756297004103

Farr ML. 1976. Flora Neotropica, Monograph No:16. N.Y. Bot. Garden.

Farr ML. 1981. How to know the true slime molds. Wm. C. Brown Company Publishers. lowa USA.

190 ... Oran & Ergiil

Giinal N. 1997. Tiirkiyede Baslica Agag Tirlerinin Cografi Yayilislari, Ekolojik ve Floristik
Ozellikleri. Cantay Kitabevi, Istanbul, Turkey.

Harkonen M. 1987. Some additions to the knowledge of Turkish myxomycetes. Karstenia 27: 1-7.

Harkonen M, Uotila P. 1983. Turkish myxomycetes developed in moist chamber cultures. Karstenia
de Tord be

Harkonen M, Rikkinen J, Ukkola T, Enroth J, Virtanen V, Jaaskalainen K, Rinne E, Hiltunen L,
Piippo S, He X. 2004. Corticolous myxomycetes and other cryptogams on seven native tree
species in Hunan Province, China. Systematics and Geography of Plants 74: 189-198.

Ing B. 1999. The myxomycetes of Britain and Ireland. The Richmond Publishing Co. Ltd. England.

Keller HW, Brooks, TE. 1977. Corticolous myxomycetes VII: contribution toward a monograph of
Licea, five new species. Mycologia 69: 667-684. http://dx.doi.org/10.2307/3758857

Lado C. 2001. Nomenmyx: a nomenclatural taxabase of myxomycetes. Cuadernos de Trabajo de
Flora Micoldgica Ibérica 16.

Lado C, Pando E. 1997. Flora Mycologica Iberica. Vol. 2. CSIC, Madrid, Spain.

Martin GW, Alexopoulos CJ. 1969. The myxomycetes. Univ. of Iowa Press, Iowa.

McHugh R. 1998. Corticolous myxomycetes from Glen Mhuire, Co. Wicklow. Mycologist 12:
166-168. http://dx.doi.org/10.1016/S0269-915X(98)80072-4

Mitchell DW. 2013. Inventory of all-world myxomycetes, dichotomous and synoptic keys, etc.
Privately published by the author. DVD [updated].

Nannenga-Bremekamp NE. 1991. A guide to temperate myxomycetes. Biopress Limited. Bristol.

Segmen O, Gemici Y, Gérk G, Bekat L, Leblebici E. 1995. Tohumlu Bitkiler Sistematigi. Ege
Universitesi Fen Fakiiltesi Kitaplar Serisi. No. 116. Izmir, Turkey

Sesli E, Denchev CM. 2014. Checklists of the myxomycetes, larger ascomycetes, and larger
basidiomycetes in Turkey. 6th edn. Mycotaxon Checklists Online
(http://www.mycotaxon.com/resources/checklists/sesli-v106-checklist.pdf): 1-136.

Stephenson SL. 1989. Distribution and ecology of myxomycetes in temperate forest. II. Patterns
of occurrence on bark surface of living trees, leaf litter, and dung. Mycologia 81: 608-621.
http://dx.doi.org/10.2307/3760136

Stephenson SL, Stempen, H. 1994. Myxomycetes: a handbook of slime molds. Timber Press,
Portland, Oregon USA.

Thind KS. 1977. The myxomycetes of India. ICAR, New Delhi.

MY COTAXON

http://dx.doi.org/10.5248/130.191
Volume 130, pp. 191-196 January-March 2015

A new species of Lophodermium with
variously branched paraphyses

Hal-LIn Gu’, YA-FEI Xu, DAN-DAN Lv’,
SHI-JUAN WANG’, & YING-REN LIN?*

' School of Life Science &? School of Forestry & Landscape Architecture,
Anhui Agricultural University, West Changjiang Road 130, Hefei, Anhui 230036, China

*CORRESPONDENCE TO: yingrenlin@yahoo.com

AxsstrRact — Lophodermium heterocladum, a new fungus developing on leaves of
Rhododendron cavaleriei, was collected from the Mount Sanqingshan National Park of Jiangxi
Province, China. Description, illustration, and comments are provided for this taxon. The
type collection is deposited in the Reference Collection of Forest Fungi of Anhui Agricultural
University, China (AAUF).

Key worps — foliicolous fungus, taxonomy, fungal diversity, Ericaceae

Introduction

During research on leaf-inhabiting fungi on members of the Ericaceae
in the Mount Sanqingshan National Park of China, one specimen of the
ascomycete family Rhytismataceae was obtained. Examination of that material
and comparison with previously described species (Lin et al. 2001a,b) led to
the conclusion that the specimen represents a new species of Lophodermium
Chevall. Fifty-seven Lophodermium species have previously been reported
from China (Hou et al. 2009, Luo et al. 2010, Shi et al. 2010, Chen et al. 2012,
Lin et al. 2012, Wang et al. 2012, Chen et al. 2013, Gao et al. 2013, Wang et al.
2014, Lu et al. 2015).

Materials & methods

The conidiomata and ascomata were examined macroscopically with a dissecting
microscope at 10—50x magnification. After rehydration in water for ca 10 min, fruiting
bodies were cut into 10-15 um thick sections using a freezing microtome. The sections
were mounted in water, KOH solution, Melzer’s reagent, cotton blue in water, or
lactophenol-cotton blue and examined microscopically. The colors of various structures

192 ... Gu &al.

were observed in water. Gelatinous sheaths surrounding ascospores and paraphyses
were examined in 0.1% (w/v) lactophenol-cotton blue. Measurements were made using
material in 5% KOH and from more than 30 paraphyses, asci, ascospores, conidiogenous
cells, and conidia for each specimen. Point and line integrated illustrations of external
status and internal structures of fruiting bodies were drawn using a microscopic drawing
device. The type collection is deposited in the Reference Collection of Forest Fungi of
Anhui Agricultural University, Hefei, China (AAUF).

Taxonomy

Lophodermium heterocladum H.L. Gu & Y.R. Lin, sp. nov. Figs 1-8
MycoBank MB 808959

Differs from Lophodermium implicatum by its variably shaped paraphyses that branch
1-2(-3) times, the presence of periphysoids and hyphal bridges, and a well-developed
subhymenium forming a strongly concave bowl shape.

TYPE: China, Jiangxi, Mount Sangingshan National Park, Huaiyushan, alt. ca 600 m, on
fallen leaves of Rhododendron cavaleriei H. Lév. (Ericaceae), 18 August 2012, Y.R. Lin, F.
Zhou & L. Zhang 2604 (Holotype, AAUF 68712).

Erymo oey: heterocladum, referring to variably branched paraphyses of the fungus.

ZONE LINES absent.

CoONIDIOMATA developing on upper side of leaves scattered in pale sandy
brown, subcircular or irregular, pale areas with an obvious edge. In surface
view, conidiomata 100-170 um diam., rounded or subrounded, black-
brown in the centre and the perimeter line of the conidioma, grey-brown
elsewhere, slightly raising the substratum surface, discharging spores through
an inconspicuous central ostiole. In median vertical section, conidiomata
intraepidermal, lenticular. UPPER WALL extremely poorly developed, composed
of a small amount of tiny angular cells. BASAL WALL dark brown, consisting of
1-3 layers of 3-5 um diam., thick-walled angular cells. SUBCONIDIOGENOUS
LAYER 4-10 um thick, comprised of colorless, thin-walled angular cells 1-3
um diam. TRICHOGYNES 32-38 x 2.5-3.2 um, cylindrical, tapering towards the
rounded apex, 1-3-septate in the lower half. ConrpIoPHORES not observed.
CONIDIOGENOUS CELLS 10-14 x 1.8-2.4 um, ampullaceous, colorless, usually
proliferating sympodially and percurrently. Conrip1A 4-6 x 0.8-1 um, hyaline,
aseptate, cylindrical, sometimes slightly curved, obtuse or round at the ends,
smooth-walled.

Ascomarta in similar positions to conidiomata on the host, scattered to clustered,
occasionally coalescent. In surface view, ascomata 350-680 x 170-360 um,
elliptical, ends round or obtuse, black, slightly shiny, with a clearly marked
outline, moderately raising the substratum surface, but slightly sunken
near the split, opening by a single longitudinal split which extends almost
to the ascomatal edge. Lips absent. In median vertical section, ascomata
intraepidermal. COVERING STROMA 12-20 um thick, mainly composed of

Lophodermium heterocladum sp. nov. (China) ... 193

Fics 1-7. Lophodermium heterocladum (holotype) on Rhododendron cavaleriei. 1. Habit on a leaf.
2. Detail of fruiting bodies. 3. Ascoma in median vertical section. 4. Portion of ascoma in median
vertical section. 5. Paraphyses, asci and ascospores. 6. Conidioma in vertical section. 7. Conidia.

194 ... Gu &al.

Fic. 8. Lophodermium heterocladum (holotype): variously branched paraphyses.

Lophodermium heterocladum sp. nov. (China) ... 195

textura angularis with dark brown to black, thick-walled cells 3-5 um diam.,
connecting to the basal stroma. There is a 6-8 um thick mulch which comprises
strongly carbonized tissue with no obvious cellular structure outside the top
of the covering stroma. Periphysoids lining the inner face of the covering
stroma, 5-14 x 1-1.5 um, cylindrical, not swelling at the apex, hyaline, straight
or slightly curved, 0-4-septate. BASAL STROMA dark-brown, consisting of 1-3
layers of thick-walled angular cells 3-7 um diam. The triangular space between
the covering stroma and basal stroma is filled with hyaline, thin-walled angular
cells 4-9 um diam. SUBHYMENIUM well developed, 15-22 um thick, strongly
concave and forming a bowl shape, composed of colorless textura angularis
and intricata. PARAPHYSES 130-155 x 1.5-2.2 um, filiform, hyaline, thinly
septate, mostly gradually or abruptly swollen to 4.2—-5.8 um at the apex,
branching 1-2(-3) times often near the top and in the middle or occasionally
in the positions not far from the base, sometimes with hyphal bridges near the
base between adjacent paraphyses, surrounded by a ca 0.5 um thick gelatinous
matrix. AScI ripening sequentially, 110-140 x 5.5-7.2 um, cylindrical, apex
round, short-stalked, thin-walled, J-, 8-spored. AscosporEs arranged in a
fascicle, 85-120 x 1.2-1.6 um, filiform, hyaline, aseptate, straight or slightly
curved, apex round or obtuse, hardly tapered towards the round or acute base,
with a thin gelatinous sheath.

HOST SPECIES, HABITAT, & DISTRIBUTION: Producing conidiomata and
ascomata on fallen leaves of Rhododendron cavaleriei. Known only from the
type locality, Jiangxi Province, China.

ComMENTs — Lophodermium heterocladum is very similar to L. implicatum
Y.R. Lin & Z.S. Xu on Rhododendron maculiferum subsp. anhweiense (E.H.
Wilson) D.F. Chamb. in the way ascomata and conidiomata are embedded and
in ascal shape and size. However, L. implicatum has ascomata on both surfaces
of the leaf, dark brown zone lines, the corner between the covering stroma
and basal stroma consisting of a very small amount of colorless, thin-walled
angular cells, paraphyses branched near the apex and entangled to form a light
yellow-brown epithecium 8-12 um thick, subhymenium composed of colorless
textura porrecta, and lacks periphysoids and hyphal bridges (Lin et al. 2001a).

Lophodermium rufum Y.R. Lin & K. Lion Rhododendron maculiferum subsp.
anhweiense is easily distinguished from the new species by its much larger
(620-1040 x 430-580 um) subcuticular ascomata, well-developed rufous lips,
covering and basal stroma consisting of textura angularis-epidermoidea, wider
(8—10.5 um) asci, shorter and wider (54—86 x 1.5—2 um) ascospores containing
many granules, unbranched paraphyses slightly gradually swollen at the apex
(Lin et al. 2001b).

196 ... Gu & al.

Acknowledgments

The authors are grateful to Dr J.E. Taylor and Dr M. Ye for pre-submission reviews
leading to the improvement of our manuscript, and to Mr FE Zhou and Ms L. Zhang
for the field investigations. This study was supported by the National Natural Science
Foundation of China (No. 31270065, 31170019).

Literature cited

Chen JL, Lin YR, Hou CL, Wang SJ. 2012 [“2011”]. Species of Rhytismataceae on Camellia spp.
from the Chinese mainland. Mycotaxon 118: 219-230. http://dx.doi.org/10.5248/118.219

Chen L, D.W. Minter, Wang SJ, Lin YR. 2013. Two new species of Rhytismataceae on fagaceous
trees from Anhui, China. Mycotaxon 126: 109-120. http://dx.doi.org/10.5248/126.109

Gao XM, Lin YR, Huang HY, Hou CL. 2013. A new species of Lophodermium associated with the
needle cast of Cathay silver fir. Mycological Progress 12: 141-149.
http://dx.doi.org/10.1007/s11557-012-0817-y

Hou CL, Piepenbring M. 2009. Lophodermium puerense sp. nov. on needles of Pinus yunnanensis vat.
tenuifolia from southwest Yunnan. Mycotaxon 107: 259-262. http://dx.doi.org/10.5248/107.259

Lin YR, Liu HY, Hou CL, Wang SJ, Ye M, Huang CL, Xiang Y, Yu SM. 2012. Flora fungorum
sinicorum, vol. 40, Rhytismatales [in Chinese]. Science Press, Beijing. 261p.

Lin YR, Xu ZS, Li K. 2001a. Two new species of the genus Lophodermium from the Huangshan
Mountains [in Chinese]. Mycosystema 20: 457-460.

Lin YR, Xu ZS, Li K, Xie YS. 2001b. Two new species of Lophodermium Chev. on Anhwei
rhododendron [in Chinese]. Journal of Anhui Agricultural University 28: 358-361.

Lu DD, Tang YP, Wang LH, Wang SJ, Lin YR. 2015 [“2014”]. Lophodermium quadrisporum sp.
nov. (Rhytismataceae) on Rhododendron faberi subsp. prattii. Mycotaxon 129(2): 459-463.
http://dx.doi.org/10.5248/129.459

Luo JT, Lin YR, Shi GK, Hou CL. 2010. Lophodermium on needles of conifers from Yunnan Province,
China. Mycological Progress 9:235-244. http://dx.doi.org/10.1007/s11557-009-0632-2

Shi GK, Luo JT, Hou CL. 2010. Species of the Rhytismataceae on Pinus armandii [in Chinese].
Mycosystema 29(2): 159-163.

Wang SJ, D.W. Minter, Gao XM, Lin YR. 2012. A new species of Lophodermium on Saccharum
arundinaceum [in Chinese]. Mycosystema 31(4): 471-475.

Wang SJ, Xu YF, Tang YP, Li Q, Lin YR. 2014. Lophodermium urniforme, a new species of Rhytismataceae
from China. Mycosystema 33(4): 768-772. http://dx.doi.org/10.13346/j.mycosystema. 140029

MY COTAXON

http://dx.doi.org/10.5248/130.197
Volume 130, pp. 197-205 January-March 2015

Bertia hainanensis sp. nov. (Coronophorales)
from southern China

LARISSA N. VASILYEVA ', HAI-XIA MA?,
ALEKSEY V. CHERNYSHEV>", & STEVEN L. STEPHENSON?

‘Institute of Biology & Soil Science, Far East Branch of the Russian Academy of Sciences,
Vladivostok 690022, Russia

?Institute of Tropical Bioscience & Biotechnology, Chinese Academy of Tropical Agricultural Sciences,
Haikou 571101, China

°A.V. Zhirmunsky Institute of Marine Biology, Far East Branch of the Russian Academy of Sciences,
Vladivostok 690059, Russia

‘Far Eastern Federal University, Vladivostok 6900950, Russia

*Department of Biological Sciences, University of Arkansas, Fayetteville, Arkansas 72701, USA

* CORRESPONDENCE TO: vasilyeva@biosoil.ru

ABSTRACT — A new species of Bertia collected in southern China is described and illustrated.
Bertia hainanensis is characterized by a combination of such features as non-ostiolate
ascomata with a roughened tuberculate surface, 1-septate cylindrical-geniculate ascospores,
and filiform paraphyses. It is most similar to B. tropicalis, from which it differs by its narrower
ascospores and its filiform paraphyses. The distinctive features of the ascomatal surface were
examined with a scanning electron microscope. The validity of the family Bertiaceae is
discussed.

KEY worpDs — Ascomycota, coronophoralean fungi, SEM images, taxonomy

Introduction

In 2012, the second author collected a specimen of Bertia De Not. from
the Bawangling National Nature Reserve (Hainan Province, China), and we
consider it to represent a species new to science. As in our previous studies
on several other coronophoralean fungi (Vasilyeva et al. 2010, 2012a, 2013),
we prepared images of fruiting bodies using a scanning electron microscope
(SEM).

There are only a few records of Bertia species from China. Bertia macrospora
Sacc. and B. moriformis (Tode) De Not. have been reported from Gansu, Jiangsu,

198 ... Vasilyeva & al.

Jilin, and Yunnan provinces (Teng 1996); B. sinensis J.C. Krug & Corlett was
described from Yunnan (Krug & Corlett 1988) and B. biseptata Sivan. & W.H.
Hsieh from Taiwan (Hsieh et al. 1995).

Materials & methods

The type specimen of the new species is deposited in the Fungarium of the Chinese
Academy of Tropical Agricultural Sciences, Haikou, Hainan, China (FCATAS). Material
was prepared for SEM examination in accordance with Vasilyeva et al. (2012a). Light
micrographs of asci and ascospores were obtained using a Leica DM 4500B microscope,
and a Zeiss EVO 40 scanning electron microscope was used to examine surface details
of the ascomata.

Taxonomy

Bertia hainanensis Lar.N. Vassiljeva, HX. Ma, Chernyshev &
S.L. Stephenson, sp. nov. Fie. 1
MycoBank MB 857516
Differs from Bertia tropicalis by its narrower ascospores and its filiform paraphyses.
Type: China, Hainan Province, Bawangling National Nature Reserve, on dead branches

of an unidentified tree, 5 November 2012, Hai-Xia Ma (Holotype: FCATAS 300).

EryMo_oey: refers to the province of China where the fungus was collected.

AsScOMATA superficial, mostly aggregated, turbinate, with a thick sterile base,
collapsing collabent when dry, black, 640-820 um diam., with a roughened,
tuberculate surface, tubercules 50-75 x 35-50 um.; walls 40-60 um thick,
consisting of 3-4 layers of angular cells 7-11 x 6-8 um; Munk pores present,
0.8-1 um diam. Asci elongated-clavate, 8-spored, long-stipitate, with a kind
of apical ring ca. 2 um, in the spore-bearing portions (72-)80-100 x 14-18
um, stalks 100-120 um, oozing from ascomata in fascicles intermingled with
numerous filiform paraphyses. Ascosporgs overlapping biseriate, cylindrical,
basal one-third curved, geniculate, 1-septate, hyaline, 24-28 x 4.8-5.5 um.

Notes: The most similar species described in the literature appears to be Bertia
tropicalis Huhndorf et al., which differs by its wider ascospores (5-9.5 um) and
the presence of inflated paraphyses (Huhndorf et al. 2004). Bertia tropicalis
is known from Costa Rica, French Guiana, Jamaica, Panama, and the United
States (Puerto Rico), all localities that are consistent with a Caribbean center
of fungal biodiversity. As such, B. tropicalis could be considered as a vicarious
species with respect to B. hainanensis, which might be expected to occur in
other countries within the southeastern Asian center of fungal diversity (cf.
Vasilyeva et al. 2012b). Bertia hainanensis differs from both B. tropicalis and
B. sinensis in its more narrow ascospores but shares with B. tropicalis the
collapsing collabent ascomata in contrast to the elongated and non-collapsing
ascomata that characterize B. sinensis.

Bertia hainanensis sp. nov. (China) ... 199

Fic.1. Bertia hainanensis (holotype, FCATAS 300): a-d. Ascomata; e. Ascomatal surface; f, g. Cross
section through a lateral wall; h. Fascicle of asci and paraphyses; i. Ascus with ascospores; j. Ascal
apex showing a tiny apical ring; k. Ascospores; |. Paraphyses. Scale bars: a = 0.5 mm; b, c = 200 um;
d, h = 100 um; e, i= 50 um; f, k, 1 = 20 um; g, j =5 um.

Other members of the relatively small group of Bertia species known from
southeastern Asia are B. fructicola Henn. and B. oxyspora (Penz. & Sacc.) Hohn.
from Indonesia (Hennings 1894a; Penzig & Saccardo 1897), B. novoguineensis

200 ... Vasilyeva & al.

Henn. from the northeast of the island of New Guinea (Hennings 1894b), and
B. tessellata Petch and B. turbinata Petch from Sri Lanka (Petch 1922). Almost
all of these have wider ascospores than B. hainanensis, with B. oxyspora also
characterized by longer ascospores (<50 um). Only B. tessellata has an ascospore
width (4.5-5.0 um) that is similar to that of B. hainanensis, but ascospores of
B. tessellata are pale fuscous and 3-septate.

Discussion: some remarks on the Bertiaceae

Both the genus Bertia and the family Bertiaceae Smyk are poorly investigated,
and their current composition is uncertain. Smyk (1981) proposed the family
without referring to its distinguishing characters, although she cited the
cytological studies of ascomata and asci by several French mycologists (Luc
1952, Chadefaud 1954, Parguey-Leduc 1977) as warranting segregation of Bertia
into a new family. As circumscribed by Smyk (1981), the new family included
three genera (Bertia, Calyculosphaeria Fitzp., and Rostrocoronophora Munk),
but Calyculosphaeria species are now considered to belong to Nitschkia G.H.
Otth ex P. Karst., while Rostrocoronophora has been transferred to a different
order (Diaporthales) as a synonym of Gnomonia Ces. & De Not. (Bolay 1972).
One might wonder just which characters could support the family Bertiaceae
sensu Smyk.

A search for characters in the French literature cited by Smyk (1981) also
has not revealed the possible criteria for distinguishing the family Bertiaceae.
All three publications compared B. moriformis with species from other—
sometimes very distantly related—groups, and such a comparison has surely
provided a number of differences, but if a new family is segregated within the
order Coronophorales, as in the case of the study by Smyk (1981), it should be
based upon differences among the members of this order. Luc (1952) compared
B. moriformis and Systremma natans (‘Tode) Theiss. & Syd. [= Dothidea sambuci
(Pers.) Fr., Dothideales| and listed several characters between them (e.g., asci
with double walls thickened at the apex versus asci with single and thin walls
without a thickening at the apex), but these very same characters distinguish the
roughly outlined groups Ascohymeniales and Ascoloculares (Nannfeldt 1932).
So, although those characters properly separate B. moriformis and S. natans
into higher taxonomic groups, they cannot serve as a basis for delimiting the
family Bertiaceae since different hierarchical levels of a taxonomic system are
characterized by their own characters. In addition, Luc (1952) compared the
asci of B. moriformis with those found in Chaetomium Kuntze and Melanospora
Corda (although without referring to particular species) and concluded that B.
moriformis might be an ‘intermediate’ taxon that occupies a position between
the order Dothideales and the class Haerangiomycetes as created (Falck & Falck
1947) for Melanospora and Ophiostoma Syd. & P. Syd. However, this clearly

Bertia hainanensis sp. nov. (China) ... 201

represents an example of unfairly judging the “phylogenetic” position of
B. moriformis without properly analyzing numerous groups, and such a peculiar
comparison cannot be considered as the basis for creating the Bertiaceae.

Chadefaud (1954) compared B. moriformis with another dothidealean
species, Dothidella ulmi (C.-J. Duval) G. Winter. His conclusions were similar
to those by Luc (1952), and B. moriformis was suggested as being separate from
the Dothideales, but one cannot estimate the level of differences with only two
species under comparison. Parguey-Leduc (1977) discussed different modes
of ascus formation along with the structure of ascus wall and apical ring in
pyrenomycetous fungi. She mentioned B. moriformis only once, indicating
that its dangeardium (ie., the ascus initial) is two-celled (a rare condition
for the pyrenomycetes). However, the same dangeardium was also observed
in Cordyceps militaris (L.) Fr., Epichloe typhina (Pers.) Tul & C. Tul., and
Mycosphaerella bolleana B.B. Higgins, so that feature might also represent a case
of parallelism in different fungal groups — a relatively frequent phenomenon.
Again, one cannot find any reason to warrant the Bertiaceae as a separate family.

Quite recently Huhndorf et al. (2004) accepted the Bertiaceae because two
species (B. moriformis and B. tropicalis) form a well-supported clade on the
molecular tree of the Coronophorales. However, two circumstances should be
taken into consideration. First, a taxon cannot exist without delineating the
differences that distinguish it from other taxa at the same hierarchical level.
Second, the level of ‘clades’ (familial, subfamilial, etc.) is unknown without an
estimation of differences between tentatively segregated groups. As has been
pointed out, taxa occupying the same taxonomic level are characterized by a
state combination of the same characters (Vasilyeva & Stephenson 2012, 2013).
Still more recently, Mugambi & Huhndorf (2010) provided the necessary
distinguishing characters for the Bertiaceae, but once again these appear to
be unsatisfactory. Members of this family are supposed to have ascomata that
are mostly tuberculate (if smooth, then the ascospores are uniformly brown)
and ascospores longer than 15 um. However, ascospore size cannot be used
for discriminating among families, since this character is universally used
for species delimitation. As for the presence of tubercules, these structures
characterize many members of the Coronophorales and differ only in size and
shape (Vasilyeva et al., 2010, 2012a, 2013). Moreover, the absence of tubercules
is also permitted in the Bertiaceae, so this character clearly cannot be a criterion
for the differentiation of the family.

In general, the simple accumulation of differences for the circumscription of
a group is often useless, since it is not the number of characters but the weight
of each character that actually matters. Very often, a single difference might be
of great value. As pointed out elsewhere (Vasilyeva & Stephenson 2012), the
weight of a character is its position in the taxonomic hierarchy, and this weight

202 ... Vasilyeva & al.

depends upon the frequency at which a character is used to delimit groups at
a certain level. Judging from the characters that have been proposed for the
segregation of the Bertiaceae, we have no choice but to abandon this family
until more substantial characters are found. The position of Bertia within
Nitschkiaceae Nannf. seems acceptable for the time being, although there is a
question about the differences between Bertia and Nitschkia.

For example, Nitschkia macrospora Teng has rather large and rough tubercules
on the stromatal surface (Vasilyeva et al. 2010), a condition almost comparable
with that found in B. latispora (Corlett & J.C. Krug) Lar.N. Vassiljeva (Fic. 2).
Moreover, N. macrospora ascospores are large (30-45 um long), so this species
appears to fit the concept of Bertia, at least sensu Mugambi & Huhndorf (2010).
On the other hand, B. orbis Mugambi & Huhndorf (Mugambi & Huhndorf
2010: Fig. 5r) has ascospores that are typical for Nitschkia. The clustering of
B. orbis with other Bertia species apart from Nitschkia species in the molecular
tree (Mugambi & Huhndorf 2010: Fig. 1) is not sufficient evidence supporting
their independence. In general, the dichotomous structure of molecular trees
strongly influences the composition of clades.

To illustrate this situation, we can use one of the ascomycetous families. If one
keeps the combinatorial variation of fungi in mind and attempts to construct
a dichotomous tree of the genera of the family Gnomoniaceae, the type genus
Gnomonia will fall in the same clade as Apiognomonia, Gnomoniella, and
Ophiognomonia when ascomatal type is used to define this clade. In contrast,
Gnomonia will appear in the same clade as Chalcosphaeria, Melanopelta, and
Plagiostoma when ascospore type found in each genus is considered (Vasilyeva
& Stephenson 2010). Molecular data might support both arrangements (if
they are correlated with defining features) or point towards new ones, but it
is highly probable that all genera in the Gnomoniaceae comprise a network of
taxa instead of reflecting a cladistic structure, with several characters that are
of equal value defining genera in the combinatorial space. The same situation
might be observed among the genera that comprise the Coronophorales, and we
have already pointed out the partial combinatorial variation within this order
(Vasilyeva et al. 2013). The same conclusion could be reached by considering
the key to genera provided by Mugambi & Huhndorf (2010), where several
differences (e.g., asci 8-spored vs. multispored and ascomata spinulose vs. not
spinulose) are repetitive in the delimitation of genera. The choice of any such
equal-leveled characters will lead to clades with different compositions in the
dichotomous ‘phylogenetic’ trees.

An independent existence of Bertia might be warranted in part by the
presence of a peculiar kind of ascospore—they are wormlike or cylindrical and
curved geniculate, similar to those found in some species of Lasiosphaeria Ces.
& De Not., including such examples as L. hirsuta (Fr.) Ces. & De Not., L. hispida

Bertia hainanensis sp. nov. (China) ... 203

d. Ascospores. Scale bars: a = 200 um; b = 100 um; c = 50 um; d = 20 um.

(Tode) Fuckel, and L. ovina (Pers.) Ces. & De Not. Ascospore types have been
traditionally used to segregate genera in ascomycetous fungi, and we can follow
this tradition for the time being.

Acknowledgments

We are grateful to the engineer D.V. Fomin for his assistance in providing the
micrographs produced at the Far Eastern Center of Electron Microscopy of the A.V.
Zhirmunsky Institute of Marine Biology (Vladivostok). We also thank Prof. Makoto
Kakishima (Jilin Agricultural University, China) and Dr. Adam Rollins (Lincoln

204 ... Vasilyeva & al.

Memorial University, USA) for serving as presubmission reviewers and for providing
helpful comments and suggestions.

Literature cited

Bolay A. 1972 [“1971”]. Contribution 4 la connaissance de Gnomonia comari Karsten. Etude
taxonomique, phytopathologique et recherches sur sa croissance in vitro. Berichte der
Schweizerischen Botanischen Gesellschaft 81: 398-482.

Chadefaud M. 1954. Sur les asques de deux Dothideales. Bulletin de la Société Mycologique de
France 70: 99-108.

Falck R, Falck O. 1947. A new class of Ascomycetales. A contribution to the orbis vitae system of
fungi. Palestine Journal of Botany, Rehovot series, 6: 89-106.

Hennings P. 1894a. Neue und interessante Pilze aus dem K6nigl. botanischen Museum in Berlin
II. Hedwigia 33: 229-233.

Hennings P. 1894b. Fungi novo-guineenses II. Botanische Jahrbiicher fiir Systematik,
Pflanzengeschichte und Pflanzengeographie 18, Beiblatt 44: 22-40.

Hsieh WH, Chen CY, Sivanesan A. 1995. Taiwan fungi: new species and new records of ascomycetes.
Mycological Research 99: 917-931. http://dx.doi.org/10.1016/S0953-7562(09)80750-8

Huhndorf SM, Miller AN, Fernandez FA. 2004. Molecular systematics of the Coronophorales
and new species of Bertia, Lasiobertia and Nitschkia. Mycological Research 108: 1384-1398.
http://dx.doi.org/10.1017/S0953756204001273

Krug JC, Corlett M. 1988. A new species of Bertia from China. Canadian Journal of Botany 66:
1256-1258. http://dx.doi.org/10.1139/b88-179

Luc M. 1952. Structure et développement de deux Dothideales: Systremma natans (Tode) Th. et Syd.
et Bertia moriformis (Tode) de Not. Bulletin de la Société Mycologique de France 68: 149-164.

Mugambi GK, Huhndorf SM. 2010. Multigene phylogeny of the Coronophorales: morphology and
new species in the order. Mycologia 102: 185-210. http://dx.doi.org/10.3852/09-043

Nannfeldt JA. 1932. Studien uber die Morphologie und Systematik der nicht-lichenisierten
inoperculaten Discomyceten. Nova Acta Regiae Societatis Scientarum Upsaliensis, ser. 1V 8(2):
1-368.

Parguey-Leduc A. 1977. Les asques de Pyrénomycetes. Revue de Mycologie 41: 281-339.

Penzig O, Saccardo PA. 1897. Diagnoses fungorum novorum in insula Java collectorum. Malpighia
11: 387-409.

Petch T. 1922. Additions to Ceylon fungi (II). Annals of the Royal Botanic Gardens, Peradeniya
Ziad 9-322.

Smyk LV. 1981. Fungi of the order Coronophorales in the mycoflora of the Ukraine. Ukrainian
Journal of Botany 38(6): 46-49 [in Ukrainian].

Teng SC. 1996. Fungi of China. Mycotaxon, Ltd., Ithaca. 586 p.

Vasilyeva LN, Stephenson SL. 2010. The problems of traditional and phylogenetic taxonomy of
fungi. Mycosphere 1: 45-51.

Vasilyeva LN, Stephenson SL. 2012. The hierarchy and combinatorial space of characters in
evolutionary systematics. Botanica Pacifica 1: 21-30.

Vasilyeva LN, Stephenson SL. 2013. An essentialistic view of the species problem. 141-169, in: IY
Pavlinov (ed.). The species problem. Ongoing issues. Rijeka: InTech.

Vasilyeva LN, Chernyshev A, Stephenson SL. 2010. Pyrenomycetes of the Russian Far
East 4: family Nitschkiaceae (Coronophorales, Ascomycota). Mycologia 102: 233-247.
http://dx.doi.org/10.3852/09-090

Bertia hainanensis sp. nov. (China) ... 205

Vasilyeva LN, Chernyshev AV, Stephenson SL. 2012a. Neochaetosphaerella thaxteriospora gen. et
sp. nov. and Tympanopsis texensis sp. nov. (Coronophorales, Ascomycota) from Texas, USA.
Fungal Diversity 52: 191-196. http://dx.doi.org/10.1007/s13225-011-0124-y

Vasilyeva LN, Stephenson SL, Hyde KD, Bakhali AH. 2012b. Some stromatic pyrenomycetous
fungi from northern Thailand - 1. Biscogniauxia, Camillea and Hypoxylon (Xylariaceae).
Fungal Diversity 52: 65-76. http://dx.doi.org/10.1007/s13225-011-0150-9

Vasilyeva LN, Chernyshev AV, Stevenson SL, Hyde KD. 2013. Tortulomyces thailandicus gen. et sp.
nov. and Nitschkia siamensis sp. nov. (Coronophorales, Ascomycota) from northern Thailand.
Mycoscience 54: 110-115. http://dx.doi.org/10.1016/j.myc.2012.09.004

MYCOTAXON

http://dx.doi.org/10.5248/130.207
Volume 130, pp. 207-213 January-March 2015

Distribution of Alternaria species among sections. 1.
Section Porri

PHILIPP B. GANNIBAL

Laboratory of Mycology and Phytopathology, All-Russian Institute of Plant Protection,
Shosse Podbelskogo 3, Saint Petersburg, 196608, Russia

CORRESPONDENCE TO: phbgannibal@yandex.ru

ABSTRACT — The evaluation of all available descriptions of Alternaria species allowed the
assignment of another 71 species to A. sect. Porri, in addition to the 46 species previously
included on the basis of phylogenetic analysis. We assess that this section encompasses 117
species, which are listed. An emended and somewhat expanded description of A. sect. Porri
is presented.

Key worps — Nimbya, N. rhapontici

Introduction

The genus Alternaria Nees is a large taxonomic group that comprises
approximately 280 species (Simmons 2007). A series of large-scale works
has attempted to resolve the phylogeny of Alternaria and other alternarioid
hyphomycetes by employing more than ten different genomic loci (Pryor &
Bigelow 2003; Hong et al. 2005; Runa et al. 2009; Lawrence et al. 2012, 2013,
2014; Woudenberg et al. 2013). Several well-supported phylogenetic lineages
have been revealed within the alternarioid hyphomycetes that have led to
several taxonomic novelties. The genus Alternaria was divided into eight
taxonomic sections by Lawrence et al. (2013). According to an alternative
concept (Woudenberg et al. 2013) all alternarioid hyphomycetes were collapsed
into one genus, Alternaria, with 24 sections, 11 of which referred to Alternaria
sensu stricto.

The monophyletic status of the so-called large-spored species was supported
by all analyses. Both research groups (Lawrence et al. 2013; Woudenberg et
al. 2013) agreed that the group of large-spored species should be treated as a
well-defined section of the genus, Alternaria sect. Porri, based on phylogenetic
as well as morphological data. Forty-six species were included in the section

208 ... Gannibal

by Lawrence et al. (2013). However, the size of the section was estimated to
comprise c. 120 (Gannibal 2012) or c. 80 (Woudenberg et al. 2013) species.
Alternaria porri was chosen as type species. Only one large-spored species,
A. helianthiinficiens E.G. Simmons et al., was placed apart from section Porri
(Woudenberg et al. 2013).

The main discriminating morphological features of many Alternaria sect.
Porri species are larger conidia and the presence of a long filiform apical beak
(Lawrence et al. 2013, Woudenberg et al. 2013). Sexual morphs have not yet
been discovered. Probably all species in this section are parasites of plants.
Species of A. sect. Porri have a rather strong substrate specialization and usually
infect either a single plant species or only few species within a single family.

It is unlikely that all 280 Alternaria species will be molecularly characterized
in the foreseeable future. Moreover, for several species, living isolates are not
known and herbarium material is mostly too old and scanty for successful
DNA extractions. On the other hand, the morphological diversity of Alternaria
is well studied and available in a monograph published by Simmons (2007).
Classical comparative morphology can be utilized to establish the affiliation of
all Alternaria species to a specific section.

The aim of this work was to detect among phylogenetically unexamined
Alternaria species, as far as possible, all species that fit the current morphological
circumscription of A. sect. Porri and to emend the morphological concept of
this section to reflect its extended membership.

Materials & methods

Morphology of almost all Alternaria species with legitimate names was analyzed
with regard to conformity with criteria of Alternaria sect. Porri. The morphological
assessment was based predominantly on descriptions made by Simmons (2007).
Species described after 2007 were assessed using original diagnoses and illustrations.
Descriptions of eight species included in A. sect. Porri were found in Sun & Zhang 2007
(A. lactucicola and A. rhaponticicola); Labuda et al. 2008 (A. jesenskae); Zhang & Zhang
2008 (A. amphicarpaeae, A. sojae, and A. tribuli); and Gannibal 2010 (A. simmonsii
and A. silybi). Two conceptions of A. sect. Porri were used for comparison, those of
Lawrence et al. (2013) and Woudenberg et al. (2013).

Results & discussion

The evaluation of all available descriptions of Alternaria species allowed for
the addition of 71 species to A. sect. Porri. The size of this section is assessed to
comprise 117 species. Alternaria species of A. sect. Porri are listed in TABLE 1.

Eight species for which living specimens are not available—A. bannaensis
W.Q. Chen & TY. Zhang, A. basellae T.Y. Zhang, A. calystegiae Nelen, A. ellisii
Pandotra & Ganguly, A. iridicola (Ellis & Everh.) J.A. Elliott, A. nicotiana J.L.
Cheng, A. nyctanthis K.B. Deshp. & Rajd., and A. trachelospermicola (T.Y. Zhang

Alternaria sect. Porri...

TABLE 1. List of Alternaria spp. assigned to A. sect. Porri.

> PPB BP BS

BPPeeeeereaereeererreeeeeePer eee e eer eszee eee eee eee

acalyphae (Nelen) E.G. Simmons [a]

. acalyphicola E.G. Simmons (Lawrence et al. 2013)

. africana E.G. Simmons [b]

. agerati Sawada ex E.G. Simmons (Lawrence et al. 2013)

. agripestis E.G. Simmons & K. Mort. (Lawrence et al. 2013)

_ allii Nolla

. amphicarpaeae Meng Zhang & T.Y. Zhang

. anagallidis A. Raabe (Lawrence et al. 2013)

. anodae E.G. Simmons

. aragakii E.G. Simmons (Lawrence et al. 2013)

. argyroxiphii E.G. Simmons & Aragaki (Lawrence et al. 2013)
. ascaloniae E.G. Simmons & C.F. Hill

. azadirachtae E.G. Simmons & Alcorn

. bataticola Ikata ex W. Yamam. (de Hoog & Horre 2002; Lawrence et al. 2013)
. beringelae E.G. Simmons

. beticola E.G. Simmons & C.F. Hill

. blumeae E.G. Simmons & Sontirat (Hong et al. 2005; Lawrence et al. 2013)
. bonducellae R. Dubey et al. [b]

. brasiliensis .M. Queiroz et al.

. calendulae Ondiej (Lawrence et al. 2013)

. capsici E.G. Simmons (Hong et al. 2005; Lawrence et al. 2013)
. caricae T.Y. Zhang et al. [b]

. carthami S. Chowdhury (Lawrence et al. 2013)

. cassiae Jurair & A. Khan (Lawrence et al. 2013)

. catharanthi (T.Y. Zhang & X.E. Lin) E.G. Simmons [b]

. centaureae E.G. Simmons

. cepulicola V.G. Rao [b]

. ciceris E.G. Simmons

. cichorii Nattrass (Lawrence et al. 2013)

. cirsinoxia E.G. Simmons & K. Mort. (Lawrence et al. 2013)

. crassa (Sacc.) Rands (Pryor & Bigelow 2003; Hong et al. 2005; Lawrence et al. 2013)
. cretica E.G. Simmons & Vakal. (Lawrence et al. 2013)

. cucumericola E.G. Simmons & C.F. Hill

cucumerina (Ellis & Everh.) J.A. Elliott (Hong et al. 2005; Lawrence et al. 2013)

. cyamopsidis Rangaswami & A.V. Rao

. cyphomandrae E.G. Simmons (Lawrence et al. 2013)

. danida E.G. Simmons (Lawrence et al. 2013)

. daturicola T.Y. Zhang et al. [b]

. dauci (J.G. Kiihn) J.W. Groves & Skolko (Pryor & Bigelow 2003; Hong et al. 2005;

Lawrence et al. 2013; Woudenberg et al. 2013)

. dichondrae Gambogi et al. (Lawrence et al. 2013)

. echinaceae E.G. Simmons & C.F. Hill

. eichhorniae Nag Raj & Ponnappa

. enydrae S.A. Khan & M. Kamal [b]

. euphorbiae (Barthol.) Aragaki & J.Y. Uchida [b]

. euphorbiicola E.G. Simmons & Engelhard (Hong et al. 2005; Lawrence et al. 2013)

A. flagelloidea (G.F. Atk.) Luttr. [b]
A. gaurae E.G. Simmons & C.F. Hill

209

210 ... Gannibal

A. glyceriae E.G. Simmons & C.F. Hill

A. grandis E.G. Simmons (Lawrence et al. 2013)

A. guangxiensis W.Q. Chen & T.Y. Zhang [b]

. hawaiiensis E.G. Simmons (Lawrence et al. 2013)

. heliophytonis E.G. Simmons

. herbiculinae E.G. Simmons

. hibiscinficiens E.G. Simmons & C.F. Hill

. hordeiseminis E.G. Simmons & G.F. Laundon

. hortensiae Unamuno ex E.G. Simmons [b]

. iranica E.G. Simmons & Ghosta

. jesenskae Labuda et al.

. lactucicola X. Sun & T.Y. Zhang

. lallemantiae (Khokhr.) E.G. Simmons [c]

. latispora T.Y. Zhang & Meng Zhang [b]

. limicola E.G. Simmons & M.E. Palm (Lawrence et al. 2013)

. linariae (Neerg.) E.G. Simmons

. linicola J.W. Groves & Skolko (de Hoog & Horre 2002; Lawrence et al. 2013)

. longirostrata T.Y. Zhang & Meng Zhang [b]

. loofahae E.G. Simmons & Aragaki

. lunariae (Oudem. & C.J.J. Hall) E.G. Simmons [b]

. macrospora Zimm. (Pryor & Bigelow 2003; Hong et al. 2005; Lawrence et al. 2013;
Woudenberg et al. 2013)

. montanica E.G. Simmons & Robeson

. multirostrata E.G. Simmons & C.R. Jacks. (Lawrence et al. 2013)

. nattrassii E.G. Simmons [b]

. nitrimali E.G. Simmons & M.E. Palm (Lawrence et al. 2013)

. novae-guineensis E.G. Simmons & C.F. Hill

. obtecta E.G. Simmons

A. passiflorae J.H. Simmonds (Lawrence et al. 2013)

A. petasitis M. Kubota et al. [b]

A. pipionipisi E.G. Simmons

A. poonensis Ragunath (Lawrence et al. 2013)

A. porri (Ellis) Cif. (Pryor & Bigelow 2003; Hong et al. 2005; Lawrence et al. 2013;

Woudenberg et al. 2013)
A. protenta E.G. Simmons (Lawrence et al. 2013)
A. pseudorostrata E.G. Simmons (Hong et al. 2005; Lawrence et al. 2013; Woudenberg et
al. 2013)

A. pulcherrimae T.Y. Zhang & J.C. David

. ranunculi E.G. Simmons

. readeri (G. Winter) Neerg. [b]

. rhaponticicola X. Sun & T.Y. Zhang

. ricini (Yoshii) Hansf. (Lawrence et al. 2013)

. rosa-sinensis M.X. Gao & T.Y. Zhang [b]

. rosifolii E.G. Simmons & C.F. Hill

. rostellata E.G. Simmons (Lawrence et al. 2013)

. saposhnikoviae J.Z. Zhang & T.Y. Zhang [b]

. sauropodis E.G. Simmons

. scorzonerae (Aderh.) Loer. (Lawrence et al. 2013)

. sesami (E. Kawam.) Mohanty & Behera (de Hoog & Horre 2002; Lawrence et al. 2013)

SPP PP PPR PPR RBBB eB

> P PPB Se BBP BB

Alternaria sect. Porri... 211

. sidae E.G. Simmons

. silybi Gannibal

. simmonsii Gannibal

. sojae Meng Zhang & TY. Zhang

. solani Sorauer (Pryor & Bigelow 2003; Hong et al. 2005; Lawrence et al. 2013;
Woudenberg et al. 2013)

. solani-nigri R. Dubey et al. (Lawrence et al. 2013)

. spinaciae Allesch. & F. Noack [b]

. stachytarpheticola E.G. Simmons [b]

. steviae Ishiba et al. (Lawrence et al. 2013)

. subcylindrica E.G. Simmons & R.G. Roberts (Lawrence et al. 2013)

. subtropica E.G. Simmons [b]

. sudanensis E.G. Simmons [b]

. tabasco E.G. Simmons & R.G. Roberts

. tagetica S.K. Shome & Mustafee (Lawrence et al. 2013; Woudenberg et al. 2013)

. thunbergiae E.G. Simmons & Alcorn

. tillandsiae E.G. Simmons & C.F. Hill

. tomato (Cooke) L.R. Jones [d]

. tomatophila E.G. Simmons (Lawrence et al. 2013)

. tribuli Meng Zhang & T.Y. Zhang

. tropica E.G. Simmons (Lawrence et al. 2013)

. vanuatuensis E.G. Simmons & C.F. Hill

. venezuelensis E.G. Simmons & Rumbos

. viciae-fabae E.G. Simmons & G.F, Laundon

. zinniae M.B. Ellis (Lawrence et al. 2013)

> > SB DB

SPPPPP PRR PEEP PBB eeBeB

Bold font names = species with phylogenetic data (in the cited references).
Unannotated names = species with reliable living cultures and herbarium specimens.
Names with square bracketed annotations = species with herbarium specimens as the only
available reliable material:

[a] = representative isolates have not been chosen;.

[b] = no known living isolates;

[c] = isolates are non-sporulating;

[d] = isolates are not correctly identified.

et al.) E.G. Simmons—were found to have intermediate characteristics that do
not allow unambiguous placement in A. sect. Porri or any other section. Dubious
morphological traits and the absence of isolates for detailed morphological or
molecular studies make it impossible to assess their phylogenetic positions
within the genus.

An isolate of A. tomato (CBS 114.35, deposited in 1935) clustered with strong
support in the autonymous clade, Alternaria sect. Alternaria (Lawrence et al.
2013). However, the credible species description in Simmons (2007) suggests
that this species resides in A. sect. Porri. According to Simmons (2007), no
correctly identified A. tomato isolates are known. Hence, we suggest that
A. tomato sensu Simmons belongs in A. sect. Porri.

Phylogenetic data revealed the polyphyletic nature of the genus Nimbya E.G.
Simmons (Lawrence et al. 2012). Only four studied species from Cyperaceae

212 ... Gannibal

and Juncaceae clustered together as a separate clade (“true” Nimbya) when three
species from Amaranthaceae were placed in Alternaria sect. Alternantherae
(Lawrence et al. 2012, 2013). There are five Nimbya species found on plants
other than Cyperaceae, Juncaceae, and Amaranthaceae, which likely belong
to other clades. They have relatively large conidia and some of them could be
related to the group of large-spored Alternaria species. At least N. rhapontici
(Nelen) E.G. Simmons is morphologically very similar to Alternaria sect.
Porri, but additional examinations are necessary to clarify the phylogeny and
taxonomy of this species.

Here we present an emended and somewhat expanded description of the
section:

Alternaria sect. Porri D.P. Lawr., Gannibal, Peever & B.M. Pryor, Mycologia 105:
541. 2013; emend. Gannibal.

On V-8 primary conidiophores short, moderately long (up to 150-250 um)
or rarely very long (up to 700 um or probably even longer [A. pseudorostrata,
A. nitrimali]), simple or branched, with one or few (up to 9 on branched
conidiophore [A. solani]) conidiogenous loci. Conidia solitary, in short
or rarely moderately long chains (up to 7 units on PCA [A. pseudorostrata,
A. euphorbiicola]), simple or with solitary lateral conidia, body of mature
conidia moderately large (40-110 x 11-25(-30) um), broadly ovoid, obclavate,
ellipsoid, subcylindrical, or obovoid, disto- and euseptate, slightly constricted at
septa, number of transepta in the conidial body varying from 7-9 [A. cirsinoxia]
to 12-19 [A. crassa, A. grandis], with a single septum or 2-3 longitudinal
septa in one or almost all transverse divisions; conidia of some species almost
always without any longitudinal septa [A. agripestis, A. subcylindrica], pale
yellowish tan to dark olivaceous brown, smooth or punctulate, at least some
mature conidia with terminal beaks, solitary or in groups (up to 5) appearing as
several beaks or a single beak with several branches, beaks aseptate or septate,
50-300(-460) um long and (4—)5-6(-8) um wide near the base, apex tapering
to 1.5-3 um or giving rise in secondary conidiophores. Conidia may form
lateral or apical secondary conidiophores without filiform portion, sometimes
with one or only few lateral secondary conidiophores arising from the body or beak.

In comparison with V-8, on PCA conidia are somewhat larger and have
beaks with larger average numbers of branches (Simmons 2007; Gannibal et
al. 2014).

Acknowledgments

It is my pleasure to acknowledge the attention of Dr. Uwe Braun and Dr. Daniel
Lawrence for their presubmission reviews of this article. This work was supported by
Russian Science Foundation (project #14-26-00067).

Alternaria sect. Porri... 213

Literature cited

Gannibal PB. 2010. Taxonomic studies of Alternaria from Russia: new species on Asteraceae.
Mycotaxon 114: 109-114. http://dx.doi.org/10.5248/114.109

Gannibal PB. 2012. Chapter 11. Understanding the phylogeny of the alternarioid hyphomycetes:
what can the consequences be in taxonomy? 303-331, in: JK Misra et al. (eds). Systematics and
Evolution of Fungi. Science Publishers Inc., Enfield, New Hampshire, USA.

Gannibal PB, Orina AS, Mironenko NV, Levitin MM. 2014. Differentiation of the closely related
species, Alternaria solani and A. tomatophila, by molecular and morphological features and
aggressiveness. Eur. J. Plant Pathol. 139: 609-623. http://dx.doi.org/10.1007/s10658-014-0417-6

Hong SG, Cramer RA, Lawrence CB, Pryor BM. 2005. Alt a 1 allergen homologs from Alternaria
and related taxa: analysis of phylogenetic content and secondary structure. Fungal Genet. Biol.
42: 119-129. http://dx.doi.org/10.1016/j.fgb.2004.10.009

de Hoog GS, Horré R. 2002. Molecular taxonomy of the Alternaria and Ulocladium species
from humans and their identification in the routine laboratory. Mycoses 45: 259-276.
http://dx.doi.org/10.1046/j.1439-0507.2002.00747.x

Labuda R, Elias P Jr, Sert H, Sterflinger K. 2008. Alternaria jesenskae sp. nov., a new species
from Slovakia on Fumana procumbens (Cistaceae). Microbiol. Res. 163: 208-214.
http://dx.doi.org/10.1016/j.micres.2006.05.004

Lawrence DP, Park MS, Pryor BM. 2012. Nimbya and Embellisia revisited, with nov.
comb. for Alternaria celosiae and A. perpunctulata. Mycol. Progress 11: 799-815.
http://dx.doi.org/10.1007/s11557-011-0793-7

Lawrence DP, Gannibal PB, Peever TL, Pryor BM. 2013. ‘The sections of Alternaria: formalizing
species-group concepts. Mycologia. 105(3): 530-546. http://dx.doi.org/10.3852/12-249

Lawrence DP, Gannibal PB, Dugan FM, Pryor BM. 2014. Characterization of Alternaria
isolates from the infectoria species-group and a new taxon from Arrhenatherum,
Pseudoalternaria arrhenatheria sp. nov. Mycol. Progress 13(2): 257-276.
http://dx.doi.org/10.1007/s11557-013-0910-x

Pryor BM, Bigelow DM. 2003. Molecular characterization of Embellisia and Nimbya species and
their relationship to Alternaria, Ulocladium and Stemphylium. Mycologia 95: 1141-1154.
http://dx.doi.org/10.2307/3761916

Runa F, Park MS, Pryor BM. 2009. Ulocladium systematics revisited: phylogeny and taxonomic
status. Mycol. Progress 8: 35-47. http://dx.doi.org/10.1007/s11557-008-0576-y

Simmons EG. 2007. Alternaria. An identification manual. Utrecht: CBS. 775 pp.

Sun X, Zhang T-Y. 2007. Taxonomic studies of Alternaria 10: Two new species and a new record
from China. Mycotaxon 101: 283-287.

Woudenberg JHC, Groenewald JZ, Binder M, Crous PW. 2013. Alternaria redefined. Studies in
Mycology. 75: 171-212. http://dx.doi.org/10.3114/sim0015

Zhang M, Zhang T-Y. 2008. Taxonomic studies of Alternaria from China 11. Three large-spored
new species. Mycotaxon. 103: 263-268.

MY COTAXON

http://dx.doi.org/10.5248/130.215
Volume 130, pp. 215-221 January-March 2015

Morchella galilaea, an autumn species from Turkey

HatTirA TASKIN™, Hasan HUseyIn DoGAn?, & SAADET BUYUKALACA'

‘Department of Horticulture, Faculty of Agriculture, University of Cukurova,
01330 Adana, Turkey

*Department of Biology, Faculty of Science, University of Selcuk
42074, Konya, Turkey

*CORRESPONDENCE TO: hatirataskin1@gmail.com

Asstract — Morchella specimens collected during the autumn seasons of 2009-12 from
Adana Province, Turkey, have been identified by DNA sequencing as representatives of
Morchella galilaea. The macro- and micromorphology, phylogeny, ecology, and fruiting
phenology of the Turkish material are evaluated.

KEY worps — mist units, morels, taxonomy, volcanic media

Introduction

Morchella species (true morels) are some of the most valuable and important
fungi consumed worldwide due to their unique aroma and flavor. Their
commercial value has been increasing over the years as a result of growing
consumer demand. Countries with diverse morel species include China, USA,
Pakistan, India, and Turkey, which export morels to European countries (Pilz
et al. 2007). In Turkey, morels generally grow in pine forests (Pinus brutia,
P. nigra) from late February to June (Taskin & Bityiikalaca 2012). According to
a review by Pilz et al. (2007), researchers have reported Morchella associated
with elm, ash, aspen, tulip poplar, apple, and cottonwood trees. Although morel
fruiting season occurs generally during the spring season, Sturgis (1905) and
Masaphy et al. (2009) have reported morels fruiting in different seasons (Pilz et
al. 2007). Morels may be found throughout Turkey, especially in Mediterranean
and Aegean regions. If they can be collected in relatively large volumes, morels
offer the potential of providing a significant income for people living near the
forests and could be regarded as a significant export product for Turkey.

In addition to the existence of an endemic Turkish morel species,
M. anatolica \siloglu et al. (Isiloglu et al. 2010), Taskin et al. (2010, 2012)

216 ... Taskin, Dogan, & Bityiikalaca

identified 20 phylogenetically distinct Morchella species in Turkey. Nineteen
of these species were collected from forests in spring, but one (given the
phylogenetic code Mes-16) was observed in autumn in the mist units of the
Horticultural Research and Application Area of Cukurova University, Adana,
Turkey (Taskin et al. 2012). Based on studies by Taskin et al. (2012), O'Donnell
et al. (2011), Du et al. (2012a,b) we named this population as a new species
but later realized that it represented Morchella galilaea, described from Israel
by Clowez (2012). However, as detailed morphological, microscopical, and
ecological features were not provided in the original description, we have
evaluated the morphology, phylogenetic position, ecology, and fruiting
phenology of M. galilaea collected from Turkey. Our results should be beneficial
to future studies concerned with culture and conservation of Morchella spp.

Materials & methods

Specimens were collected from Adana province in Turkey (Horticultural Research
and Application Area of Cukurova University) during autumn and early winter
(October-December) between 2009 and 2012 (Fic. 1). Two (in 2009), two (in 2010),
and 11 (in 2012) Morchella specimens were collected from mist units used for rooting
agricultural plant cuttings.

Molecular analyses were performed at the United States Department of Agriculture
(USDA-NCAUR-Peoria-IL) in USA (Taskin et al. 2012). These samples were analyzed
through partial RNA polymerase I (RPB1), RNA polymerase II (RPB2), translation
elongation factor (EFl-a), 28S (LSU) rRNA gene and internal transcribed spacer
(ITS) rRNA gene sequences. Macro- and microscopical characters were studied at
the Department of Biology, Selcuk University (Konya, Turkey). The specimens were
mounted in Melzer’s reagent and examined microscopically using a Leica DM 750
microscope. Volcanic tuff was analyzed at Alata Horticultural Research Institute, and
pH (Jackson 1962), salinity (Soil Survey Division Staff 1993), total lime (CaCO,; Caglar
1949), organic matter (Walkley & Black 1934), phosphorus (P; Olsen & Dean 1965),
magnesium (Mg), calcium (Ca), iron (Fe), zinc (Zn), manganese (Mn), and copper (Cu)
were analyzed using an atomic absorption spectrophotometer (Varian SpectrAA Model
220FS, Germany).

Taxonomy

Morchella galilaea Masaphy & Clowez, Bull. Soc. Mycol. Fr. 126 (3-4): 238.
2012 [“2010”]. Fics 1-2
ASCOMATA 53-78 mm tall. HymMENopHORE 36-45 mm tall, 12.5-17.5
mm wide, conic to subcylindric, pitted and ridged, ridges have two types,
these are narrow polygonal ridges and long elliptical ridges. STERILE RiBs
longitudinally arranged, 5.2-9.8 mm wide, 14.5-24.74 mm tall, thin, elastic,
simple, or forked and anastomosing to form elongated hymenial pits, white,
bluntly rounded when young, but often flattened in age, ridges glabrous.

Morchella galilaea in Turkey ... 217

Fic. 1 - Morchella galilaea specimens collected from mist units
on the Cukurova University campus (Adana, Turkey).

HyMENIAL Pits light gray, whitish gray, blackish gray or silvery gray, when
young; becoming, greyish-light brown, light olive brown or yellow brown with
maturity. ASCOCARP MARGIN connecting directly into the stalk without a sharp
bend, free. Stipe 17-33 mm high, 5-8.2 mm wide, cylindrical and thickening
slightly towards the base; surface when young completely glabrous and white,
in maturity light yellowish white. CoNTExT whitish, 0.5-1 mm thick in the
hollow hymenophore; sterile inner surface whitish, glabrous. AscosPORES
elliptical, smooth (surface distinctly rugulate to tectate under the scanning
electron microscope), with homogeneous contents, (12.5-)13.5-18(-19.5)
x (7.5-)8-9.5(-10) um [average Q =1.83]. Asci 8-spored, 165-220 x 15-22
um, cylindric, hyaline. PARAPHYSES cylindric to subclavate, apices rounded to
subacute, with 2-3-septate, hyaline or with brown to brownish homogeneous
contents in KOH (2%), 100-130 x 6-11 um. RESIDUAL PARAPHYSES on sterile
ridges similar to paraphyses, hyaline or with bundled brownish contents.
SPECIMENS COLLECTED: Turkey. Adana: Cukurova University Campus, 37°01’55”N
35°22'12”E, alt. 39 m, solitary or scattered on volcanic tuff in misted cutting propagation

beds, 17 Nov 2009, leg. H. Taskin, Taskin 117 (ANK); 28 Nov 2012, leg. H.H. Dogan,
Dogan 10022 (Kon Fungarium).

218 ... Taskin, Dogan, & Bityiikalaca

EHT=20.00kKV ——Signal A= SE1 Mag= 6.00 KX
WD = 10.5 mm IProbe= 20pA

-
7

Fic. 2 - Morchella galilaea (Turkish specimen, Taskin 117).
A, B: asci with ascospores. C: paraphyses. D: ascospore (SEM).

Discussion

The morphological features of M. galilaea were cited by Clowez (2012) as:
ascomata 3-5 cm height, ovoid, light yellow; primary alveolus large, few; edges
of ribs same color and rusty; stipe cylindrical rather slender, attenuated at the
top and wider at the base.

When we compared M. galilaea samples collected from Israel and Turkey
morphologically, we found certain differences between the samples, which
we summarize as follows: (a) ascomata of the Israel specimens were shorter
(3-5 cm); (b) the stipes of the Turkish specimens were more or less cylindrical
except for the slightly thicker base while in the Israeli collection the stipes had
a narrower apex and wider base; (c) the Israeli specimen was found under
Fraxinus syriaca, while our Cukurova University specimens were collected
from a no tree zone.

Morchella galilaea in Turkey ... 219

Molecular characterization

Morchella galilaea [as Mes-16] has been molecularly assessed through
GCPSR analyses by Taskin et al. (2012) and also molecularly analyzed by
O’Donnell et al. (2011), Du et al. (2012 a, b), Masaphy et al. (2010), Clowez
(2012), and Richard et al. (2015). These studies indicate that M. galilaea occurs
in China (Du et al. 2012a,b), Java, Hawaii, Israel (Masaphy et al. 2010, Clowez
2012), New Zealand, India (Kanwal et al. 2011), and three African countries
as well as in Turkey. The Turkish and Israel sequences of Morchella galilaea
are deposited in GenBank-ITS1-5.8S-ITS2 as JQ723082 and GU589858,
respectively (Masaphy et al. 2010; Clowez 2012; Du et al. 2012 a, b).

Ecological assessment

The results of the volcanic tuff analyses are provided in TABLE 1. Relatively
high levels of copper, manganese, zinc, iron, magnesium and calcium were
detected in the volcanic tuff medium, which was identified as having a loam
soil structure, normal CaCO,, and alkali salinity profile. There was a relatively
low potassium level. Singh et al. (2004), who determined that pH values vary
from 6.5 to 7 in morel natural areas in India, detected high levels of calcium,
carbon, nitrogen, sodium, and lead and low levels of phosphates, chloride, and
potassium; soil type varied from sandy to loam.

Sturgis (1905) observed morels fruiting in September in an aspen-spruce
forest burned the previous summer in southwestern British Columbia (Pilz
et al. 2007). Masaphy et al. (2009) reported that a Morchella species emerged
from late autumn to late spring in northern Israel under Rhamnus alaternus,

TABLE 1. Properties of the volcanic tuff in which Morchella galilaea was found fruiting.

PROPERTY RESULTS
Texture (100 g/ml) 46
Caco, (%) 19.6
Salinity (mmhos/cm) 0.06
Organic matter (%) 2.6
pH 7.6
Available potassium (ppm) 167.5
Available phosphorus (ppm) 19.2
Calcium (ppm) 2300
Available magnesium (ppm) 759
Iron (ppm) 6.6
Zinc (ppm) 2.8
Manganese (ppm) 4.8

Copper (ppm) 1.6

220 ... Taskin, Dogan, & Bityiikalaca

Platanus orientalis, Populus euphratica, Laurus nobilis, and Fraxinus syriaca.
However, they could not find a mycorrhizal relationship between trees and
morels. Morchella is known to be saprobic. However, according to a review study
conducted by Tedersoo et al. (2010), Morchella species may associate weakly
with plant roots and thus are likely to be facultative biotrophs. Nonetheless,
some researchers believe that certain Morchella species may be able to form
mycorrhizal relationships with certain trees. Dahlstrom et al. (2000) reported
a mycorrhizal relationship between Morchella and species of Pinaceae, and
Buscot & Kottke (1990) detected a mycorrhizal association between Morchella
and Picea abies. Our morel specimens grew in a mist unit. While this unit was
used for rooting olive cuttings during 2009-2010, no cuttings were placed
into the units in 2012, which suggests that our specimens are probably not
mycorrhizal. Our hypothesis is that the mycelium of the species was carried
to the mist units by means of the volcanic tuff, where it developed over the
years. After mycelial development, a sprinkle irrigation system and hormone
application used for rooting might also have stimulated Morchella fruiting. The
same species has been found under Populus and Quercus by Du et al. (2012b)
and under Fraxinus syriaca by Masaphy et al. (2010). Although morels typically
fruit from late February to early June, our Morchella specimens were collected
between October and December. In 2012, the air temperature at Cukurova
University-Adana averaged 29°C in October, 23°C in November, and 16°C in
December.

Acknowledgements

The authors would like to express their sincere gratitude to Dr. Kerry O'Donnell
(USDA-NCAUR- Peoria, IL, USA) for allowing us to perform molecular analyses in his
laboratory. A special thanks goes to Dr. Davut Keles (Manager of Alata Horticultural
Research Institute), for the volcanic tuff analysis, and to Berken Cimen, for organization
of the figures. We are also indebted to Prof. Dr. Mitko Karadelev, Prof. Dr. Aysun Peksen,
Prof. Dr. Daniel Royse for their critical reviews of the manuscript.

Literature cited

Buscot F, Kottke I. 1990. The association of Morchella rotunda (Pers.) Boudier with roots of Picea
abies (L.) Karst. New Phytologist 116: 425-430.

Caglar KO. 1949. Soil science. Ankara University Agriculture Faculty Publication, Ankara.

Clowez P. 2012 (‘2010’). Les morilles: une nouvelle approche mondiale du genera Morchella.
Bulletin de la Société Mycologique de France 126: 199-376.

Dahlstrom JL, Smith JE, Weber NS. 2000. Mycorrhiza-like interaction by Morchella with species of
the Pinaceae in pure culture synthesis. Mycorrhiza 9: 279-285.

Du XH, Zhao Q, Yang ZL, Hansen K, Taskin H, Bitytikalaca S$, Dewberry D, Moncalvo JM, Douhan
GW, Robert V, Crous PW, Rehner SA, Rooney AP, O'Donnell K, Sink S. 2012a. How well do
ITS rDNA sequences ‘barcode’ species of true morels (Morchella)? Mycologia 104: 1351-1368.
http://dx.doi.org/10.3852/12-056

Morchella galilaea in Turkey... 221

Du XH, Zhao Q, Yang ZL, O'Donnell K, Rooney AP, Yang ZL. 2012b. Multigene molecular
phylogenetics reveals true morels (Morchella) are especially species-rich in China. Fungal
Genetics and Biology 49: 455-469. http://dx.doi.org/10.1016/j.fgb.2012.03.006

Isiloglu M, Alli H, Spooner BM, Solak MH. 2010. Morchella anatolica (Ascomycota), a new species
from southwestern Anatolia, Turkey. Mycologia 102: 455-458. http://dx.doi.org/10.3852/09-186

Jackson ML. 1962. Soil chemical analysis. Prentice Hall, New York.

Kanwal HK, Acharya K, Ramesh G, Reddy MS. 2011. Molecular characterization of Morchella
species from the western Himalayan region of India. Current Microbiology 62: 1245-1252.
http://dx.doi.org/10.1007/s00284-010-9849-1

Masaphy S, Zabari L, Goldberg D. 2009. New long season ecotype of Morchella rufobrunnea from
northern Israel. Micologia Aplicada International 21: 45-55.

Masaphy S, Zabari L, Goldberg D, Jander-Shagug G. 2010. The complexity of Morchella systematics:
a case of the yellow morel from Israel. Fungi 3(2): 14-18.

O’Donnell K, Rooney AJ, Mills GL, Kuo M, Weber NS, Rehner SA. 2011. Phylogeny and historical
biogeography of true morels (Morchella) reveals an early Cretaceous origin and high continental
endemism and provincialism in the Holarctic. Fungal Genetics and Biology 48: 252-265.
http://dx.doi.org/10.1016/j.fgb.2010.09.006

Olsen SR, Dean LA. 1965. Estimation of available phosphorus in soils by extraction with sodium
bicarbonate. United States Department of Agriculture, Washington, D.C.

Pilz D, Mclain R, Alexander S, Villarreal-Ruiz L, Berch S, Wurtz TL, Parks CG, McFarlane E, Baker
B, Molina R, Smith JE. 2007. Ecology and management of morels harvested from the forests of
western North America. General Technical Report, PNW-GTR-710. United States Department
of Agriculture, Forest Service, Pacific Northwest Research Station, Portland.

Richard F, Sauve M, Bellanger JM, Clowez P, Kansen H, O'Donnell K, Urban A, Courtecuisse
R, Moreau PA. 2015. True morels (Morchella, Pezizales) of Europe and North America:
Evolutionary relationships inferred from multilocus data and a unified taxonomy. Mycologia
http://dx.doi.org/10.3852/14-166.

Singh SK, Kamal S, Tiwari M, Rai RD, Upadhyay RC. 2004. Myco-ecological studies of natural morel
bearing sites in Shivalik Hills of Himachal Pradesh, India. Micologia Aplicada International 16:
1-6.

Soil Survey Division Staff. 1993. Soil survey manual. United States Department of Agriculture
Handbook 18. Soil Conservation Service.

Sturgis WC. 1905. Remarkable occurrence of Morchella esculenta (L.) Pers. Journal of Mycology
Like2.69;

Taskin H. Biyitikalaca S. 2012. Morel (Morchella) mushroom. Bah¢e 41: 25-36.

Taskin H, Buyiikalaca $, Dogan HH, Rehner SA, O’Donnell K. 2010. A multigene molecular
phylogenetic assessment of true morels (Morchella) in Turkey. Fungal Genetics and Biology 47:
672-682. http://dx.doi.org/1016/j.fgb.2010.05.004

Taskin H, Bityitikalaca S, Hansen K, O’Donnell K. 2012. Multilocus phylogenetic analysis of true
morels (Morchella) reveals high levels of endemics in Turkey relative to other regions of Europe.
Mycologia 104: 446-461. http://dx.doi.org/10.3852/11-180

Tedersoo L, May TW, Smith M. 2010. Ectomycorrhizal lifestyle in fungi: global diversity,
distribution, and evolution of phylogenetic lineages. Mycorrhiza 20: 217-263.

Walkley A, Black LA. 1934. An examination of the Degtjareff method for determining soil organic
matter and a proposed modification of the chromic acid titration method. Soil Science 39:
29-38.

MY COTAXON

http://dx.doi.org/10.5248/130.223
Volume 130, pp. 223-225 January-March 2015

Hyphoderma hallenbergii,
a new corticioid species from India

MANINDER Kaur, AVNEET P. SINGH’, & G.S. DHINGRA

Department of Botany, Punjabi University, Patiala 147002, India

*CORRESPONDENCE TO: avneetbot@gmail.com

ABSTRACT — Hyphoderma hallenbergii sp. nov. from Himachal Pradesh, India is described.

Key worps — Baghi, Basidiomycota, Agaricomycetes, Meruliaceae

During August 2012 fungal forays conducted of the Baghi area of district
Shimla, Himachal Pradesh, India, Maninder and Avneet collected a fungus
on decaying wood of Cedrus deodara in a mixed forest. Comparison of
macroscopic and microscopic characters (Eriksson & Ryvarden 1975, Dhingra
1989, Larsson 1998, Bernicchia & Gorjén 2010, Singh et al. 2010, Priyanka
& Dhingra 2012, Mycobank 2014) suggested that it belonged in the genus
Hyphoderma (Meruliaceae, Agaricomycetes, Basidiomycota). The fungus, which
is similar to H. nemorale K.H. Larss. and H. incrustatum K.H. Larss. in having
two types of cystidia but which differs in basidiospore size, is described here as
a new species.

Hyphoderma hallenbergii Man. Kaur, Avneet P. Singh & Dhingra, sp.nov. Plate 1
MycoBank MB 809801
Differs from Hyphoderma nemorale and H. incrustatum by its smaller and ellipsoid

basidiospores.

Type: India, Himachal Pradesh: Shimla, Narkanda, on Baghi road, on the bark of
decaying wood of Cedrus deodara (Roxb. ex D. Don) G. Don, 19 August 2012, Maninder
Kaur & Avneet 6962 (PUN, holotype).

Erymo.oey: In the honor of Dr. Nils Hallenberg, Professor Emeritus, University of
Gothenburg, Sweden.

Basidiocarp resupinate, adnate, effused, <100 um thick in section; hymenial
surface smooth to somewhat grandinioid, chalky white; margins thinning

224 ... Kaur, Singh, & Dhingra

9, aS yo > SIN
a8. g

ae
ye

PiatTE 1. Hyphoderma hallenbergii (holotype). 1. Basidiocarp showing hymenial surface.
2. Capitate hymenial cystidia. 3. Basidium. 4,5. Basidiospores.
6. Vertical section through basidiocarp.

out or indeterminate, concolorous or paler. Hyphal system monomitic.
Generative hyphae <2.5 um wide, branched, septate, clamped; subiculum
poorly developed, formed of few hyphae running more or less parallel to
the substrate; subhymenial hyphae well developed, vertical, denser. Cystidia
of two types: [1] Capitate cystidia 25-44 x 5-6 um, subfusiform, originating
from the hymenium, thin-walled, with basal clamp, sparsely encrusted; [2]
Cystidia which are cylindrical to flexuous with constrictions, sometimes
almost moniliform 60-80 x 5-7 um, originating from subhymenium, slightly
thick-walled, with basal clamp, often encrusted with crystals. Basidia 16-34 x
4-5 um, clavate to subclavate, constricted to sinuous, 4-sterigmate, with basal
clamp; sterigmata <3.5 um long. Basidiospores 6.3-7.5 x 3.5-4.5 um, ellipsoid,
with oily contents, thin-walled, inamyloid, acyanophilous.

REMARKS — Hyphoderma nemorale and H. incrustatum both differ from
H. hallenbergii by their larger, cylindrical basidiospores (H. nemorale, 10-14 x
4-5 um; H. incrustatum, 11-16 x 4-5.5 um; Larsson 1998).

Hyphoderma hallenbergii sp. nov. (India) ... 225

Acknowledgements

The authors thank Head (Department of Botany, Punjabi University, Patiala) for
providing research facilities, Dr. Nils Hallenberg (Professor Emeritus, Denmark)
for expert comments and peer review; and Prof. B.M. Sharma (Department of Plant
Pathology, COA, CSKHPAU, Palampur, H.P., India) for peer review.

Literature cited

Bernicchia A, Gorjon SP. 2010. Corticiaceae s.1. Fungi Europaei 12. Edizioni Candusso. Alassio.
Italia. 1008 p.

Dhingra GS. 1989. Genus Hyphoderma Wallr. em. Donk in the Eastern Himalayas. 197-212, in: ML
Triveda et al. (eds). Plant Science Research in India. Today & Tomorrow’s Printers & Publishers,
New Delhi.

Eriksson J, Ryvarden L. 1975. The Corticiaceae of North Europe- II. Fungiflora, Oslo. pp. 288-546.

Larsson KH. 1998. Two new species in Hyphoderma. Nordic Journal of Botany. 18(1): 121-127.
http://dx.doi.org/10.1111/j.1756-1051.1998.tb01106.x

MycoBank. 2014. Fungal databases. Nomenclature and species banks. [Accessed: 16/07/2014]
http://www.mycobank.org/.

Priyanka, Dhingra GS. 2012. Two new species of Hyphoderma (Agaricomycetes) from India.
Mycotaxon 119: 255-260. http://dx.doi.org/10.5248/119.255

Singh AP, Priyanka, Dhingra GS, Singla N. 2010. A new species of Hyphoderma (Basidiomycetes)
from India. Mycotaxon 111: 71-74. http://dx.doi.org/10.5248/111.71

MY COTAXON

http://dx.doi.org/10.5248/130.227
Volume 130, pp. 227-231 January-March 2015

Xylaria thailandica — a new species from
southern Thailand

NATTHAPACH SRIHANANT™, VASUN PETCHARAT?’,
& LARISSA N. VASILYEVA?

‘Tropical Agricultural Resource Management Programme & Department of Pest Management,
Faculty of Natural Resources, Prince of Songkla University,
Hatyai, Songkhla, 90112, Thailand

*Institute of Biology and Soil Sciences, Far East Branch of the Russian Academy of Sciences,
Vladivostok 690022, Russia

* CORRESPONDENCE TO: srihanant.n@gmail.com

ABSTRACT —A new species, Xylaria thailandica, was collected from dead wood and fallen
branches in southern Thailand. This fungus has upright, gregarious, cylindrical stromata that
arise from a tomentose stipe and which is internally dark brown to black at the center but
yellow around the perithecia. Its ascospores are inequilateral and with a germ slit that spirals
lengthwise.

KEY worpDs — ascomycetes, Xylariaceae, decaying wood, fallen branches

Introduction

Fungi belonging in the genus Xylaria are saprophytic, endophytic, parasitic
(Whalley 1985), or symbionts in insect nests (Hawksworth et al. 1995, Pélaez et
al. 2008). They are characterized by upright, sessile, or stipitate stromata with
cylindrical, clavate, or irregularly shaped fertile parts. Many species have been
reported from different substrata throughout the world (Rogers & Samuels
1986, Bi et al. 1993, Rogers et al. 2005, Ju & Hsieh 2007, Ju et al. 2009, Rogers et
al. 2008, Carmona et al. 2009, Hladki & Romero 2010, Fan & Lin 2011, Fournier
et al. 2011, Rogers & Ju 2012).

Thailand is considered one area containing a high percentage of Xylaria
species, and several studies have reported on their biodiversity (Carroll 1963,
Schumacher 1982, Thienhirun 1997, Ruksawong & Flegal 2001, Chansrikul et
al. 2008, Petcharat et al. 2012). As currently recognized, Xylaria is represented

228 ... Srihanant, Petcharat, & Vasilyeva

in Thailand by 42 species. One recent collection that could not be identified
as any known Xylaria species is described and illustrated herein as Xylaria
thailandica.

Materials & methods

The specimens were collected from a natural plant site in a tropical rain forest in
southern Thailand. Hand-cut sections of dried material revived in 70% ethyl alcohol
were examined microscopically. Ascospore measurements were made from squash
mounts in lactophenol and the dimensions were determined from 20 fully mature
spores. A drop of Melzer’s reagent was added to determine the amyloid iodine reaction.
The photographs of stromata were taken using a Canon 550D DLSR digital camera.
Cultures were obtained from ascospores using the single spore isolation technique and
characterized on Potato Dextrose Agar (PDA) at room temperature (28-30 °C). All the
specimens are kept at Department of Pest Management, Faculty of Natural Resources,
Prince of Songkla University, Hatyai, Songkhla, Thailand (PSU-PM).

Taxonomy

Xylaria thailandica Srihanant, Petcharat & Lar.N. Vassiljeva, sp. nov. Fie. 1
MycoBank MB 807720
Differs from other species of Xylaria by its yellow-colored flesh, ascospores with a
spiraling germ slit, and asci with an urn-shaped apical ring.
Type— Thailand, Nakorn si thammarat Province, para rubber plantation, on dead wood

and fallen branches, 26. X. 2010, Natthapach Srihanant, (holotype PSU-PM 1059).

EryMoLocy—Refers to Thailand, where this fungus was collected.

STROMATA upright, gregarious, cylindrical, unbranched or (sometimes)
branched at the stipe, with short to long acute sterile apices, 3-7 cm total length
x 0.1-0.2 cm diam, externally at first brownish, becoming dark brown, then
black; internally dark brown to black at core, but yellow around perithecia,
texture soft, surface roughened by remnants of peeling layer and short dark
brown hairs; hairs not evident in old material. Young stroma filiform, dark
brown, but dull green at the top where the fruiting bodies produce conidia
(asexual spores). STIPE short to long, filiform, with a broadened discoid base,
covered with short brown hairs. PERITHECIA completely immersed, globose
to spherical, 210-310 um high x 135-250 um diam, ostioles papillate. Asc1
1.5-2 x 1-1.5 um, apical ring urn-shaped, blue in Melzer’s. AscosporEs brown,
unicellular, inequilateral, with broadly or narrowly rounded ends, 9-14.5 x
3-4.8 um, with spiraling germ slit spore-length.

Colonies on PDA: Colonies growth 14 days at room temperature (28-30 °C)
on PDA 9 cm diam., mycelium cottony, white with black color on the surface of
the agar. Stromatic structures not forming on media.

Xylaria thailandica sp. nov. (Thailand) ... 229

Fic. 1. Xylaria thailandica (holotype). A. Mature teleomorphic stroma on dead wood. B. Stromata
surface with conspicuous perithecial mounds. C. Transverse section of a stroma with dark color at
the core. D. Ascospores in lactophenol. E. Ascospore in Melzer’s reagent with an urn-shaped apical
ring. F Ascospore in lactophenol with spiral germ slit. G. Colonies on PDA in a 5 cm petri dish at
10 days. Scale bars: A =2 cm; B, C= 1 mm;D=10 um; E=5 um; F= 10 um; G= 1 ¢m

ADDITIONAL SPECIMENS EXAMINED—THAILAND, SONGKHLA PROVINCE, Prince
of Songkla University, Khao khohong nature trail, on fallen branches 5.XII.2010,
Natthapach Srihanant (PSU-PM 1060); SURATTHANI PROVINCE, oil palm plantation,
on dead wood in 24. I. 2011, Natthapach Srihanant (PSU-PM 1061).
ComMMENTS—Xylaria thailandica differs from other species on wood (Fournier
et al. 2011, Hladki & Romero 2010, Ju et al. 2009, Rogers & Samuels 1986,
Roger et al. 2008) by its yellow flesh and ascospores with spiraling germ slits.
This species can be separated from X. longipes var. tropica F. San Martin &
J.D. Rogers (Thienhirun 1997) and X. longipes Nitschke (Rogers et al. 2008),
which have larger ascospores and asci with a differently shaped apical apparatus.
The stromatal morphology of X. thailandica is close to X. cinerea J. Fournier
& M. Stadler (Fournier et al. 2011), which differs by its larger ascospores with
a slightly sinuous germ slit.
Comparisons between X. thailandica and other species are shown in TABLE 1.

230 ... Srihanant, Petcharat, & Vasilyeva

TABLE 1. Comparison of some wood-inhabiting species of Xylaria with the new taxon
X. thailandica.

SPECIES SPORE SIZE (um) GERMINATION SLIT APICAL APPARATUS

X. thailandica 9-14.5 x 3-4.8 Spiral, full spore-length Urn-shaped

X. apiculata 16-30 x 5-9 Straight, full spore-length Minute

X. castorea 8-14 x 4.5-6.5 Straight, probably fulllength § Minute

X. cinerea 13-17 x 5-6 Straight to slightly sinuous Tubular to urn-shaped

X. grammica 12-14.5 x 4-6.5 Straight, full spore-length Rectangular

X. hypoxylon 10-15.5 x 4.5-6.5 Straight to undulate, less Wedge-shaped
than full length

X. karsticola 11-14.5 x 4.2-6 Conspicuous straight, full Tubular with a slightly
spore length flared apex

X. longipes 9-15 x 5-6 Spiral, around spore Not seen

X. longipes var. tropica 14-16.5 x 5-6 Spiral, full spore-length Quadrate to rectangular

X. multiplex 9.5-11 x 4.5-5 Straight, almost full spore Not seen

length at the ventral side

X. zealandica 22-31 x 7.5-9 Straight to undulate, slightly | Urn-shaped
less than spore length

Acknowledgments

This work was supported by the Higher Education Research Promotion and National
Research University Project of Thailand, Office of the Higher Education Commission.
The first author would like to acknowledge Department of Pest Management, Prince of
Songkla University, providing necessary facilities in this research. We are thankful to
Dr. Julia Checa (Universidad de Alcala, Madrid, Spain) and Dr. Chaninun Pornsuriya
(Prince of Songkla University, Songkhla, Thailand) for reviewing the manuscript of our
paper and for their valuable suggestions.

Literature cited

Bi ZS, Zheng GY, Li TH. 1993. The macrofungus flora of China’s Guangdong province. Chinese
University of Hong Kong, Hong Kong.

Carmona A, Fournier J, Williams C, and Piepenbring M. 2009. New records of Xylariaceae from
Panama. North American Fungi 4(3): 1-11. http://dx.doi.org/10.2509/naf2009.004.003

Carroll GC. 1963. Studies in the flora of Thailand. XXIV. Pyrenomycetes. Dansk Botanisk Arkiv
23(1): 101-113.

Chandrasrikul A, Suwanarit P, Sangwanit U, Morinaga T, Nishizawa Y, Murakami Y. 2008. Diversity
of mushroom and macrofungi in Thailand. Kasetsart University Press, Bangkok, 514 p.

Fan ZY, Lin G. 2011. Xylaria hainanensis sp. nov. (Xylariaceae) from China. Mycosystema 30(4):
526-528.

Fournier J, Fressa F, Persoh D, Stadler M. 2011. Three new Xylaria species from southwestern
Europe. Mycological Progress 10: 33-52. http://dx.doi.org/10.1007/s11557-010-0671-8

Hawksworth DL, Kirk PM, Sutton BC, Pegler DN. 1995. Dictionary of the Fungi. 8 ed. Cambridge,
Cambridge University Press. 616 p.

Xylaria thailandica sp. nov. (Thailand) ... 231

Hladki AI, Romero AI. 2010. A preliminary account of Xylaria in the Tucuman Province,
Argentina, with a key to the known species from the Northern Provinces. Fungal Diversity 42:
79-96. http://dx.doi.org/1007/s13225-009-0008-6

Ju YM, Hsieh HM. 2007. Xylaria species associated with nests of Odontotermes formosanus in
Taiwan. Mycologia 99(6): 936-957. http://dx.doi.org/10.3852/mycologia.99.6.936

Ju YM, Vasilyeva L, Akulov A. 2009. Three new Xylaria species from Russian Far East. Mycologia
101(4): 548-553. http://dx.doi.org/10.3852/08-188

Ma HX, Vasilyeva L, Li Y. 2011. A new species of Xylaria from China. Mycotaxon 116: 151-155.
http://dx.doi.org/10.5248/116.151

Petcharat V, Worrapattamasri K, Srihanant N, Seepheuak P. 2012. Mushrooms of Southern Thailand.
Department of Pest Management, Faculty of Natural Resources, Prince of Songkla University,
Thailand. 507 p.

Pélaez F, Gonzalez V, Platas G, Sanchez-Ballesteros J, Rubio V. 2008. Molecular phylogenic studies
within the family Xylariaceae based on ribosomal DNA sequences. Fungal Diversity 31:
111-134.

Rogers JD, Samuels GJ. 1986. Ascomycetes of New Zealand 8. Xylaria. New Zealand Journal of
Botany 24(4): 615-650. http://dx.doi.org/10.1080/0028825X.1986.10409947

Rogers JD, Ju YM, Lehmann J. 2005. Some Xylaria species on termite nests. Mycologia 97(4):
914-923

Rogers JD, Miller AN, Vasilyeva LN. 2008. Pyrenomycetes of the Great Smoky Mountains National
Park. VI. Kretzschmaria, Nemania, Rosellinia and Xylaria (Xylariaceae). Fungal Diversity 29:
107-116.

Rogers JD, Ju YM. 2012. The Xylariaceae of the Hawaiian Islands. North American Fungi 7(9):
1-35. http://dx.doi.org/10.2509/naf2012.007.009

Ruksawong P, Flegal TW. 2001. Thai mushroom and other fungi. National Science and Technology
Development Agency, Thailand. 268 p.

Schumacher T. 1982. Ascomycetes from Northern Thailand. Nordic Journal of Botany 2: 257-263.

Thienhirun S. 1997. A preliminary account of the Xylariaceae of Thailand. Ph.D. Thesis. Liverpool
John Moores University. UK. 355 p.

Whalley AJS. (1985). The Xylariaceae: some ecological considerations. Sydowia 38: 369-382.

MYCOTAXON

http://dx.doi.org/10.5248/130.233
Volume 130, pp. 233-236 January-March 2015

Rosellinia brunneola sp. nov. and
R. beccariana new to China

WEI LI & LIN Guo*

State Key Laboratory of Mycology, Institute of Microbiology, Chinese Academy of Sciences, Beijing
100101, China

* CORRESPONDENCE TO: guol@im.ac.cn

ABSTRACT — A new species, Rosellinia brunneola, is described from Hainan Province, China;
and Rosellinia beccariana from Yunnan Province is described as new to China.

KEY worDs —Ascomycota, pyrenomycetous fungi, Xylariaceae, taxonomy

Rosellinia species, which are cosmopolitan in their distribution, are especially
rich in tropical and subtropical regions. 142 species have been recognized
(Petrini 2013). During mycological expeditions to tropical regions of Hainan
and Yunnan Provinces, many specimens of Rosellinia have been collected, from
which a new species and a new Chinese record are described here.

Rosellinia brunneola Wei Li bis & L. Guo, sp. nov. Fics 1-3
FUNGAL NAME FEN 570115

Differs from Rosellinia mastoidiformis by its smaller stromata and ascospores with a
straight germ slit that is as long as spore length or almost so.

Type: China, Hainan, Lingao, Juliu, alt. 250 m, on dead stems, 4.XII.2010, L. Guo 11546
(HMAS 269893, holotype).

Erymo.oey: The epithet refers to the color of ascospores.

Stromata gregarious or solitary, 0.44-0.7 mm high, 0.6-0.8 mm _ wide,
semiglobose or coniform, with top tapering to a point, surface smooth, black.
Ostioles papillate. Subiculum not seen. Ectostroma 30-60 um thick, black.
Entostroma white or brownish. Asci eight-spored, with spores arranged in
biseriate manner, cylindrical, the spore-bearing part 100-120 x 10-12.5 um,
with apical ring bluing in Melzer’s iodine reagent, cylindrical, 6-7 um high,
3-4 um broad. Ascospores brownish, asymmetrically ellipsoidal, with rounded

234 ... Li & Guo

Rosellinia brunneola sp. nov. (China) ... 235

ends, 31-39 x 5-7 um (33.8 + 3.2 x 5.9 + 1.1 um; 95%CI: 32-35 um, 5.5-6.5
um), with straight germ slit, 26-34 um long.

ComMENTs: Rosellinia brunneola is similar to R. mastoidiformis Saccas, which
differs by having larger stromata (0.5-0.9 mm high, 0.6-1 mm wide) and
ascospores with straight and indistinct germ slit that is shorter than spore
length (Petrini 2013).

Rosellinia beccariana Ces., Atti Accad. Sci. Fis. Mat. Napoli 5(21): 12. 1872.

Figs 4-6
Stromata gregarious or solitary, subglobose, sometimes with a broad, short
stipe, copper brown or black, 2.25-3.7 mm high, 2.9-3.75 mm wide, surface
smooth. Ostioles up to 600 um, pointed. Ectostroma 150-200 um thick, black.
Perithecia subglobose, 1.7-3.15 mm high, 2.6-3.5 mm wide. Asci eight-spored,
with spores arranged in biseriate manner, cylindrical, 278-355 um total length,
10-12.5(-15) um broad, the spore bearing part 201-248 um long, with apical
ring bluing in Melzer’s iodine reagent, arrowhead-shaped, 25-27 um high,
5-8 um broad. Ascospores brownish or brown, unicellular, asymmetrically
fusiform, with thread-like ends, 48-68 x 6-9 um, with straight germ slit shorter
than spore-length on flat side.

ComMMENTs: The ends of the ascospores in the Chinese material lack the slimy
sheath, which is absent in old material (Petrini 2013). The large stromata with
markedly acute ostioles make this fungus easy to recognize.

SPECIMEN EXAMINED: CHINA, YUNNAN, Mengla, Menglun, Xishuangbanna Tropical

Botanical Garden, Chinese Academy of Sciences, alt. 570 m, on corticated wood,
15.X.2013, W. Li 2903 (HMAS 253034).

Including the two species reported in this paper, 36 Rosellinia species have
been recorded in China (Teng 1963, Tai 1979, Ju & Rogers 1990, 1999, Yuan
& Zhao 1993, Liu et al. 2010, Petrini 2013).

Acknowledgements

The authors would like to express their deep thanks to Dr. L. Vasilyeva (Vladivostok,
Russia) and L.E. Petrini (Breganzona, Switzerland) for serving as pre-submission
reviewers, to Dr. Shaun Pennycook (Auckland, New Zealand) for nomenclatural review,
and to the support by the Ministry of Science and Technology of the People’s Republic
of China (No. 2013FY110400).

Fics 1-3. Rosellinia brunneola (HMAS 269893, holotype). 1. Stromata on dead stems (bar = 1 cm);
2. Ascospores; 3. Ascus. Fics 4-6. Rosellinia beccariana (HMAS 253034). 4. Stromata on wood
(bar = 1 cm); 5. Ascospores; 6. Ascus.

236 ... Li & Guo

Literature cited

Ju YM, Rogers JD. 1990. Astrocystis reconsidered. Mycologia 82: 342-349.
http://dx.doi.org/10.2307/3759905

Ju YM, Rogers JD. 1999. The Xylariaceae of Taiwan (excluding Anthostomella). Mycotaxon 73:
343-440.

Liu CE, Lu T, Gao JM, Wang MQ, Lu BS. 2010. Two new Chinese records of Rosellinia. Mycosystema
29: 459-462.

Petrini LE. 2013. Rosellinia - a world monograph. Bibliotheca Mycologica 205. 410 p.

Tai FL. 1979. Sylloge Fungorum Sinicorum. Science Press, Beijing. 1527 p.

Teng SC. 1963. Fungi of China. Science Press, Beijing. 808 p.

Yuan ZQ, Zhao ZY. 1993. Studies on the genera Amphisphaerella, Coniochaeta and Rosellinia of
Xinjiang, China. Acta Mycologica Sinica 12(3): 180-186.

MYCOTAXON

http://dx.doi.org/10.5248/130.237
Volume 130, pp. 237-239 January-March 2015

Phyllachora hainanensis sp. nov. from China

Na Lru'*, LE WANG’, Gu HUANG”, & LIN GUO?

' College of Biological Engineering, Henan University of Technology,
Lianhua Street, High & New Tech Development District, Zhengzhou 450001, China

* State Key Laboratory of Mycology, Institute of Microbiology, Chinese Academy of Sciences,
No.1 Beichen West Road, Chaoyang District, Beijing 100101, China

* CORRESPONDENCE TO: liuna3456@163.com

ABSTRACT — A newspecies, Phyllachora hainanensis on Phyllanthus emblica (Phyllanthaceae),
causing tar spots is reported from Hainan Province, China. Phyllachora hainanensis differs
from other similar species in the size and shape of ascospores.

Key worps — Sordariomycetes, taxonomy

A new species of Phyllachora (Sordariomycetes) on leaves of Phyllanthus emblica
was collected from Hainan Province, China, in November 2007. Worldwide,
only P phyllanthophila and P. phyllanthophila var. egregia have previously been
reported from the host genus Phyllanthus (Saccardo & Saccardo 1905: 834,
Saccardo & Trotter 1913: 419).

Phyllachora hainanensis Na Liu & G. Huang, sp. nov. PLATE 1
FUNGAL NAME FN570110
Differs from Phyllachora phyllanthophila by its larger ascospores.

Type: China, Hainan Province, Xiawangling, alt. 800 m, on leaves of Phyllanthus
emblica L. (Phyllanthaceae), 13 XI 2007, Shuanghui He, L. Guo & Z.Y. Li 2013 (HMAS
245338, holotype).

EtyMoLoecy: From Hainan Province, where the new species was collected.

ASCOMATA epigenous, raised, shiny, black, sparse, subglobose, ellipsoidal
or irregular, approx. 370 x 150-260 um diam., immersed in the mesophyll,
usually with a single locule, ostiole conspicuous with a conical neck extending
through the host epidermis and cuticle, without periphyses. Asci arise from
the basal and lateral walls of the ascoma, upper wall to 50 um thick, composed
of epidermal cells occluded by melanised material. Asci 60-100 x 30-70 um,

238 ... Liu & al.

PiaTE 1. Phyllachora hainanensis (HMAS 245338, holotype).
1. Ascomata; 2-4. Asci and ascospores.

8-spored, clavate, obtuse at apex, long pedunculate, thin-walled at maturity,
unitunicate. Ascosporss biseriate or irregularly biseriate, 11-18 x 9-13 um,
ellipsoidal or subglobose, one-celled, hyaline, thin-walled, smooth, without a
gelatinous sheath. ANAMORPH not seen.

CoMMENTS—Subglobose ascospores are rare in Phyllachora. Only one other
species, P. yunnanensis Na Liu & L. Guo, collected on Lespedeza (Fabaceae)
from Yunnan Province of China, has been reported as having subglobose
(sometimes ellipsoidal) ascospores. Phyllachora yunnanensis differs from
P. hainanensis in its smaller (7-13.5 x 5-9 um) ascospores (Liu & Guo 2006).
Phyllachora hainanensis is the first Phyllachora species in China discovered
on a host species in the Phyllanthaceae. Most Phyllachora species reported

Phyllachora hainanensis sp. nov. (China) ... 239

in China occur on hosts in the Poaceae, Fabaceae, and Cyperaceae (‘Tai 1979,
Teng 1963, Lan et al. 2012, Cui 2009). The only other reports of Phyllachora
on Phyllanthus are Phyllachora phyllanthophila Henn. on Phyllanthus sp.
from Peru (Hennings 1904: 250) and P. phyllanthophila var. egregia Rehm on
Phyllanthus sp. from Brazil (Rehm 1907: 530). These taxa were subsequently
transferred to Catacauma as C. phyllanthophilum (Henn.) Theiss. & Syd. and
C. egregium (Rehm) Theiss. & Syd. (Theissen & Sydow 1915: 390-391), but
Catacauma is now synonymised with Phyllachora (von Arx & Miller 1954:
212) and P. phyllanthophila var. egregia is synonymised with P. phyllanthophila
(Index Fungorum 2015). The publications cited above provide different sets of
measurements that can be summarised as 35-60 x 12-20 um for asci, and 10-
14 x 5-9 um for ascospores; thus P. phyllanthophila differs from P. hainanensis
by its smaller asci and its narrower, ellipsoidal to subovoid ascospores.

Acknowledgments

The authors would like to thank Dr. Roger Shivas (Brisbane, Australia) and Guangyu
Sun (Northwest Agriculture and Forestry University) for serving as pre-submission
reviewers, Drs. Shuanghui He and Zhenying Li for collecting the specimen, Dr. Shaun
Pennycook (Auckland, New Zealand) for nomenclatural review, and Mr. Ziyu Cao
(Institute of Botany, Chinese Academy of Sciences) for identifying the host plants.
This study was supported by the National Natural Science Foundation of China (No.
31300015) and Natural Science Research Project of Henan Province (No. 132300410350).

Literature cited

von Arx JA, Miller E. 1951. Die Gattungen der amerosporen Pyrenomyceten. Beitrége zur
Kryptogamenflora der Schweiz 11(1). 434 p.

Cui JQ. 2009. Taxonomy and molecular phylogenetics of Colletotrichum and Phyllachora. Thesis
for Master Degree of Northwest Agriculture and Forestry University. 66 p.

Hennings P. 1904. Fungi amazonici II. a cl. Ernesto Ule collecti. Hedwigia 43: 242-273.

Index Fungorum. 2015. http://www.indexfungorum.org/names/Names.asp [accessed: February
2015].

Lan JQ, Lu HJ, Wu J., Li XL, Zhang T, Kong CC, Kang ZS. 2012. Three new records of Phyllachora
in China. Mycosystema 31: 639-641. http://dx.doi.org/10.13346/j.mycosystema.2012.04.021

Liu N, Guo L. 2006. A new species of Phyllachora (Phyllachorales) on Leguminosae from China.
Mycotaxon 97: 111-114.

Rehm H. 1907. Ascomycetes novi. Annales Mycologici 5: 516-546.

Saccardo PA, Saccardo D. 1905. Supplementum universale, pars VI. Hymenomycetae -
Laboulbeniomycetae. Sylloge Fungorum 17. 991 p.

Saccardo PA, Trotter A. 1913. Supplementum universale, pars IX. Ascomycetae - Deuteromycetae.
Sylloge Fungorum 22. 1612 p.

Tai FL. 1979. Sylloge Fungorum Sinicorum. Science Press, Beijing, 1527 p.

Teng SC. 1963. Fungi in China. Science Press, Beijing. 808 p.

Theissen F, Sydow H. 1915. Die Dothideales. Kritisch-systematische Originaluntersuchungen.
Annales Mycologici 13: 149-746.

MY COTAXON

http://dx.doi.org/10.5248/130.241
Volume 130, pp. 241-246 January-March 2015

Two new species of Zasmidium from Nepal

RAVINDRA NATH KHARWAR’, ARCHANA SINGH’,
RAGHVENDRA SINGH ”, & SHAMBHU KUMAR?

‘Center of Advanced Study in Botany, Banaras Hindu University, Varanasi, U.P. India 221005

?Department of Botany, School of Biological Sciences, Dr. Harisingh Gour Central University,
Sagar, M.P, India 470003

° Birbal Sahni Institute of Palaeobotany, 53, University Road, Lucknow, U.P, India 226007

* CORRESPONDENCE TO: drsinghtaxon@gmail.com

ABSTRACT — Zasmidium cassines on living leaves of Cassine glauca (Celastraceae) and
Z. fabaceicola on living leaves of Vigna unguiculata (Fabaceae) are described and illustrated
from a forest in the terai region of Nepal. The two new foliar fungi are compared with closely
related taxa.

Key worps — foliar diseases, hyphomycetes, morphotaxonomy

Introduction

During a 1993-94 survey of anamorphic foliar fungi in the forest of the terai
region of Nepal, two undescribed species of Zasmidium were discovered. These
are characterized by having planate conidial scars, verruculose superficial
hyphae, and solitary to rarely catenate and usually rough-walled conidia.
Verruculose superficial hyphae and rough-walled conidia are characters shared
by both Stenella and Zasmidium. Molecular analyses (Arzanlou et al. 2007)
have shown that the genus Stenella is polyphyletic and that its type species,
S. araguata Syd. (Sydow 1930), clusters within the Teratosphaeriaceae.
This species is characterized by pileate conidiogenous loci, while other
former Stenella species are distinguished by having planate Cercospora-like
conidiogenous loci and belong to the Mycosphaerellaceae. Arzanlou et al.
(2007) also demonstrated that Zasmidium cellare (Pers.) Fr. (Fries 1849), the
type species of the genus Zasmidium, clusters within the Mycosphaerellaceae
and agrees both morphologically and phylogenetically with Stenella-like
hyphomycetes now included in the that family. Hence, the name Zasmidium is

242 ... Kharwar & al.

available for all former Stenella species characterized by planate conidial scars
clustering in the Mycosphaerellaceae.

Materials & methods

Diseased leaves were collected and dried to make herbarium specimens and
deposited in Herbarium Cryptogamiae Indiae Orientalis, Indian Agricultural Research
Institute, New Delhi, India (HCIO). Isotypes are retained in the herbarium of the
Department of Botany, D.D.U. Gorakhpur University, Gorakhpur, India (GPU) for
further reference. Microscopic slides of fungal structures were prepared in lactophenol
and cotton-blue mixture from infected area of leaf. Observations were made with light
microscope using oil immersion (1000x). The figures of fungal structures were drawn
using a camera lucida and show all necessary details of morphology and ontogeny of
reproductive propagules.

Taxonomy

Zasmidium cassines Kharwar, Arch. Singh, Raghv. Singh &
Sham. Kumar, sp. nov. Fig 1
MycoBank MB 809138

Differs from Zasmidium elaeodendri by its verruculose superficial mycelium, shorter,
narrower and less septate conidia and conidiophores.

Type: Nepal, Chitwan, Narayanghat, on living leaves of Cassine glauca (Rottb.) Kuntze
[= Elaeodendron roxburghii Wight & Arn.] (Celastraceae), December 1993, coll. R.N.
Kharwar, (Holotype, HCIO 42337; isotype GPU 3002).

EryMo_oey: the epithet is derived from the name of the host genus.

Infection spots amphigenous, brown, circular to subcircular, 1-6 mm
diam., coalescing and spreading to cover most of the leaf surface. Colonies
hypophyllous, effuse. Mycelium internal and external, septate, branched,
verruculose, subhyaline to pale brown, up to 3 um wide. Stromata absent.
Conidiophores arising singly as lateral branches of superficial hyphae, semi-
macronematous, mononematous, erect to procumbent, straight to flexuous,
smooth-walled, brown, 8.5-60 um x 2-4.5 um. Conidiogenous cells integrated,
polyblastic, terminal or intercalary, sympodial, cylindrical, conidiogenous
loci cicatrized with slightly thickened scars. Conidia holoblastic, dry,
acropleurogenous, simple to catenate, rarely branched, cylindrical to obclavate,
straight to curved, subhyaline to olivaceous brown, verruculose, 0-6-septate,
sometimes constricted at septa, base obconicotruncate with slightly thickened
hilum, apex acute to subacute, 13.5-109 x 1.5-4um.

Only one species of Zasmidium, Z. elaeodendri (Kamal et al.) Kamal [= Stenella
elaeodendri Kamal et al.] has been previously reported to occur on Cassine
glauca (as Elaeodendron roxburghii; Kamal 2010). Zasmidium elaeodendri
differs by its lack of verruculose superficial mycelium, its longer and wider

Zasmidium spp. nov. (Nepal) ... 243

FIGURE 1. Zasmidium cassines (HCIO 42337)
a. Infection spots. b. Mycelium. c. Conidiophores. d. Conidia.
Scale bars:A= 20 mm, B = 20 um

branched conidiophores (2126 x 4.5-6 um), and its much longer and thicker
17-septate conidia (2132 x 3.5-7 um), which are colourless, smooth-walled,
and slightly curved with a conico-truncate base and a rounded apex. The
proposed taxon, therefore, merits description as a new species.

Some other cercosporoid hyphomycete genera have been reported on
Cassine: Pseudocercospora elaeodendri (G.P. Agarwal & Hasija) Deighton on
Cassine glauca [= Elaeodendron glaucum|] and Ps. paradoxa U. Braun & Bagyan.
on Cassine [= Elaeodendron] sp. (Deighton 1976, Braun & Bagyanarayana

244 ... Kharwar & al.

1999). Pseudocercospora differs from Z. cassines in the absence of thickened
scars at the conidial bases and on conidiogenous loci.

Kamal (2010: 260-261) listed an erroneous record of Scolecostigmina
tirumalensis on Cassine glauca [= Elaeodendron glaucum}], but this is based on a
record of Stigmella tirumalensis (Bagyanarayana et al. 1992).

at
if
A

ey
1 ee,

FIGURE 2. Zasmidium fabaceicola (HCIO 42338)
a. Infection spots. b. Mycelium.c. Conidiophores.d. Conidia.
Scale bars: A = 20 mm, B = 20 um

Zasmidium fabaceicola Kharwar, Arch. Singh, Raghv. Singh &
Sham. Kumar, sp. nov. Fig 2
MycoBAnk MB 809139
Differs from Zasmidium fabacearum by its well-developed stromata, and its shorter
conidia and conidiophores.

Zasmidium spp. nov. (Nepal) ... 245

Type: Nepal, Chitwan, Narayanghat, on living leaves of Vigna unguiculata (L.) Walp.
[= Dolichos biflorus L.] (Fabaceae), January 1994, coll. R.N. Kharwar (Holotype, HCIO
42338; isotype, GPU 3413).

EryMo_oey: the epithet is derived from the host family name.

Infection spots amphigenous, blackish brown, circular to irregular, 1-15 mm
diam., later coalescing to form large irregular patches, sometimes covering
the entire leaf surface. Colonies hypophyllous, effuse, brown. Mycelium
mostly external, septate, branched, verruculose, pale brown, 1.5-3.5 um wide.
Stromata substomatal to superficial, pseudoparenchymatous, dark brown,
17-19 um in diam. Conidiophores macronematous, arising singly as lateral
branches of external hyphae, or in fascicles from stromata, erect, straight to
slightly flexuous, unbranched, smooth-walled, mid brown to dark brown below
and paler towards the apex, 4-12-septate, 75-210 x 1.5-5 um. Conidiogenous
cells integrated, polyblastic, terminal or intercalary, sympodial, cylindrical,
geniculate, scars thickened. Conidia dry, acropleurogenous, simple to catenate,
unbranched, cylindrical to obclavato-cylindrical, straight to slightly curved,
brown, verruculose, 0-3-septate, sometimes constricted at septa, hilum slightly
thickened, base obconico-truncate, apex rounded to obtuse and sometimes
cicatrized, 13.5-40 x 1.5-5um.

Only one species of Zasmidium, Z. fabacearum (K. Srivastava et al.) Kamal
(2010), has been previously reported on Dolichos, although the host was not
identified to species. Zasmidium fabacearum differs from Z. fabaceicola by
its lack of stromata and by its longer conidiophores (100-310 x 2-4 um) and
conidia (12-60 x 2-6 um).

Acknowledgments

The authors thank Prof Dr Kamal, DDU Gorakhpur University, Gorakhpur, for helpful
taxonomic advice. We are grateful to Dr. Marcin Piatek and Dr Eric H.C. McKenzie for
helpful comments and corrections. We also express our deep thanks to Dr Shaun Pennycook
for nomenclatural review and Dr Lorelei L. Norvell, Editor-in-Chief, Mycotaxon, for final
acceptance of the manuscript for publication. Last but not least, Dr Archana Singh thanks
Department of Science and Technology (DST), Government of India, New Delhi for
providing financial support under Women Scientist Scheme (WOS-A).

Literature cited

Arzanlou M, Groenewald JZ, Gams W, Braun U, Shin HD, Crous PW. 2007. Phylogenetic and
morphotaxonomic revision of Ramichloridium and allied genera. Stud. Mycol. 58: 57-93.
http://dx.doi.org/10.3114/sim.2007.58.03

Bagyanarayana G, Braun U, Jagadeeswar P. 1992. Three new phytoparasitic fungi from India.
Mycotaxon 45: 105-108.

Braun U, Bagyanarayana G. 1999. Phytopathogenic micromycetes from India (II). Sydowia. 51(1):
1519,

246 ... Kharwar & al.

Deighton FC. 1976. Studies on Cercospora and allied genera. VI. Pseudocercospora Speg., Pantospora
Cif. and Cercoseptoria Petr. Mycol. Pap. 140. 168 p.

Fries EM. 1849. Summa Vegetabilium Scandinaviae. 2: 259-572.

Kamal. 2010. Cercosporoid fungi of India. Bishen Singh Mahendra Pal Singh, Deharadun, India
351 p.

Sydow H. 1930. Fungi Venezuelani. Ann. Mycol. 28(1-2): 29-224.

MY COTAXON

http://dx.doi.org/10.5248/130.247
Volume 130, pp. 247-251 January-March 2015

The lichen genus Leiorreuma in China

X1A0-Hua WANG” ?, L1-L1 XU’, & ZE-FENG JIA”

"College of Life Sciences, Liaocheng University, Liaocheng 252059, China
’ College of Life Sciences, Shandong Agricultural University, Taian 271018, China
*CORRESPONDENCE TO: Zfjia2008@163.com

ABSTRACT — Four species of Leiorreuma (Graphidaceae, Ostropales) are reported from China.
Leiorreuma crassimarginatum is new to science, and L. dilatatum and L. melanostalazans are
new to China. Descriptions accompany a key distinguishing the four Leiorreuma species in
China. All materials are conserved in HMAS-L.

Keywords — Ascomycota, lichenized fungi, Ostropomycetidae, taxonomy,

Introduction

According to the revised generic concept (Staiger 2002), Leiorreuma Eschw.
belongs to the family Graphidaceae (Ostropales, Ostropomycetidae, Ascomycota).
The lichen genus is characterized by apothecia with conspicuous sessile opened
lirellae, a carbonised proper exciple that is often laterally thin but basally well-
developed, an inspersed hymenium, and pale brown transversely septate or
muriform ascospores that are I-. Only one species, L. sericeum, has previously
been reported from China. We report here on three additional species for the
country, one of which we describe as new to science.

Materials & methods

The specimens examined are deposited in the Herbarium Mycologicum
Academiae Sinicae - Lichenes, Beijing, China (HMAS-L). A dissecting
microscope (TEcH XTS-30D with CANon 600D camera) anda light microscope
(OLtympus SZ-51) were used for the morphological and anatomical studies.
Measurements and illustrations were taken from the manual cross-sections of
fruit bodies in tap water. The lichen substances were detected and identified by
thin-layer chromatography (TLC) (Culberson & Kristensson 1970, Culberson
1972, Orange et al. 2001).

248 ... Wang, Xu, & Jia
Taxonomy

Leiorreuma crassimarginatum Z.F. Jia, sp. nov. Piet
FUNGAL NAME FN570068

Differs from Leiorreuma hypomelaenum by its production of norstictic acid and its
larger ascospores.

Type: China. Guizhou Province, Tongren City, Mt. Fanjing, Daling, 27°55’N 108°41’E,
alt. 1570 m, on bark, 5/X/2004, coll. Jiang-Chun Wei & Tao Zhang G425 (Holotype,
HMAS-L 071776).

ErymMo_oey: Latin crassus and marginatus, in reference to the thick thalline margin.

THALLUS corticolous, crustose, grey to pale white, rough, thin, tightly attached
to the substratum, without isidia and soralia. APoTHECTIA lirelliform, short
to elongate, open, simple or sometimes branched, 1-3 mm long, 0.3—0.6 mm
wide, sessile, with thick lateral thalline margin, often rounded at the ends, not
striate, scattered over the thallus; pisc opened, pale brown; PROPER EXCIPLE
laterally slightly and basally thickly carbonized, basal exciple 90-130 um
high; EPITHECIUM brownish, 2-5 um high; HYMENIUM brownish, inspersed,
85-130 um high, I-; HypoTHEcrum brown, 15-40 um high; paRAPHYSES
simple, 1—1.5 um wide, apices unbranched; asci cylindrical to clavate, 6-spored,
80-120 x 15—25 um; ascospores brown to brownish, thick-walled, ellipsoid,
transversely septate, often 7—12-locular, 35—45 x 10-15 um.

CHEMISTRY: C-, K+ yellow to brown, P+ yellow; norstictic and stictic acids
detected by TLC.

DISTRIBUTION & ECOLOGY: Known only from the type locality, Mt. Fanjing,
an important nature reserve in Guizhou Province, South China. Other lichens
growing in the surrounding area are species of Graphis, Diorygma, and
Ocellularia.

ComMENTs: Leiorreuma crassimarginatum is characterized by its sessile
lirellae with thick lateral thalline margins, opened discs, entire labia, thickened
carbonized basal exciples, the small, brown and thick-walled ascospores,
and the presence of norstictic and stictic acids. The new lichen is similar to
L. hypomelaenum, which differs in its smaller (22—38 x 7-11 um), 7—12-locular,
and thin-walled ascospores and its lack of norstictic acid (Staiger 2002).

Leiorreuma dilatatum (Vain.) Staiger, Biblioth. Lichenol. 85: 296 (2002).

THALLUS corticolous, crustose, pale white to grey, rough to smooth, thickish.
APOTHECIA lirelliform, short to elongate, open, simple or branched, 0.5—3 mm
long, 0.3-0.7 mm wide, sessile, with thin thalline margin; pisc opened, black
to brown; PROPER EXCIPLE laterally slightly and basally thickly carbonized,
basal exciple 30—45 um high; EPprrHEcIUM brownish, 2—5 um high; HYMENIUM
brownish, inspersed, 80-100 um high, I-; HypoTHECIUM brown, 10-20 um

L. crassimarginatum sp. nov. (Leiorreuma in China) ... 249

PLaTE.1. Leiorreuma crassimarginatum (holotype, HMAS-L 071776) A. Thallus with lirellae.
B. Apothecium, cross-section. C. Ascus containing 6 ascospores. Scales: A = 1 mm; B = 100 um;
C=50 um.

high; PARAPHYSES simple, 1—1.5 um wide, apices unbranched; asci cylindrical
to clavate, 8-spored; ascospores brown to brownish, ellipsoid, transversely
septate, often 4-locular, 20-30 x 6-8 um, I+ brown.
CHEMISTRY: C-—, K-, P-; no lichen substances detected by TLC.
DISTRIBUTION & ECOLOGY: Collected on Mt. Jianfengling, the largest
remaining tropical rainforest on Hainan Island, South China. Associated
lichens included species of Graphis, Hemithecium, and Lecanora.
SPECIMEN EXAMINED: CHINA. HAINAN PROVINCE, Mt. Jianfengling, alt. 180 m, on
bark, 1/X/2008. Jing Li HN081374 (HMAS-L 127475).
ComMENTs: Leiorreuma dilatatum is characterized by sessile lirellae with
thin lateral thalline margins, opened discs, a basally carbonized exciple,
small brown ascospores, and the absence of lichen substances. It is similar to

250 ... Wang, Xu, & Jia

L. sericeum, which differs by its well-developed thalline and proper margins
and its smaller ascospores (14—20 x 5—8 um, Staiger 2002; 16-18 x 5.5—7.5 um,
our specimens). This is the first report of L. dilatatum from China.

Leiorreuma melanostalazans (Leight.) A.W. Archer, Telopea 11: 75 (2005).
THALLUS corticolous, crustose, pale grey, rough to smooth, thin. APOTHECIA
lirelliform, short to elongate, open, simple or branched, 1-4 mm long,
0.3-0.5 mm wide, sessile, with thin thalline margin; pisc opened, brown;
PROPER EXCIPLE inconspicuous, laterally slightly and basally thickly carbonized,
basal exciplel10—30 um high; EpIrHEcIUM brownish, 2—5 um high; HYMENIUM
brownish, inspersed, 90-110 um high, I-; HypoTHECIUM brown, 10-15 um
high; PARAPHYSES simple, 1—1.5 um wide, apices unbranched; asci cylindrical
to clavate, 8-spored; ascospores brown to brownish, ellipsoid, transversely
septate, often 7—9-locular, 25—40 x 8-10 um, I+ brown.
CHEMISTRY: C-, K+ yellow, P+ yellow; stictic acid detected by TLC.
DISTRIBUTION & ECOLOGY: Known from two localities in the tropical
rainforest in Hainan Island, South China. Associated lichens included species
of Graphis, Hemithecium, and Phaeographis.
SPECIMENS EXAMINED: CHINA. HAINAN PROVINCE, Mt. Jianfengling, alt. 630-700
m, on bark, 1/X/2008, Jing Li HN081422 (HMAS-L 117057); 30/IX/2008, Ze-Feng Jia
HN080683 (HMAS-L 117056).
ComMENTs: Leiorreuma melanostalazans is characterized by sessile lirellae with
thin lateral thalline margins, opened discs, a basally carbonized exciple, small
brown 7-9 locular ascospores, and the presence of stictic acid. It is similar to
L. exaltatum, which differs by its lack of lichen substances (Staiger 2002). This
is the first report of L. melanostalazans from China.

Leiorreuma sericeum (Eschw.) Staiger, Biblioth. Lichenol. 85: 305 (2002).

THALLUS corticolous, crustose, pale yellow to grey, smooth, thin. APOTHECIA
lirelliform, short to elongate, open, simple or branched, 0.5-3.5 mm long,
0.2-0.3 mm wide, sessile, with thin thalline margins, pisc opened, brown
with white-grey pruina; PROPER EXCIPLE laterally thinly and basally thickly
carbonized, basal exciple 20—40 um high; EPITHECIUM brownish, 2—5 um high;
HYMENIUM brownish, inspersed, 70-90 um high, I-; HyPOTHECIUM brown,
10-15 um high; paRAPHYSES simple, 1—1.5 um wide, apices unbranched; asc1
cylindrical to clavate, 8-spored; AscosporeEs brown to brownish, ellipsoid,
transversely septate, often 4-locular, 16-18 x 5.5—7.5 um, I+ purple-brown.

CHEMISTRY: C-—, K-, P-; no lichen substances detected by TLC.

DISTRIBUTION & ECOLOGY: Distributed throughout tropical and subtropical
South China. Associated lichens included species in the Graphidaceae and
Parmeliaceae.

L. crassimarginatum sp. nov. (Leiorreuma in China) ... 251

SPECIMENS EXAMINED: CHINA. HAINAN PROVINCE, Mt. Jianfengling, alt. 980 m, 28/

VII/2009, Jing Li HN247-1 (HMAS-L 117025). GUANGXI PROVINCE, Shangsi County,

alt. 600 m, on bark, 31/XII/1997, Shou-Yu Guo 1638 (HMAS-L).
ComMENTs: Leiorreuma sericeum is characterized by sessile lirellae with thin
lateral thalline margins, opened discs, a laterally and basally carbonized exciple,
small (<18 um long) brown 4-locular ascospores, and the absence of lichen
substances. It is similar to L. dilatatum, which differs by its larger ascospores
and poorly developed thalline and proper margins (Staiger 2002). Leiorreuma
sericeum has previously been reported (as Phaeographis sericea) from Hong
Kong (Thrower 1988, Wei 1991).

Key to species of Leiorreuma known from China

1 Ascospores 4-locular; lichen substances absent ............. 0. eee ee ee eee eee 2
1 Ascospores >6-locular; lichen substances present ........... 0... esse eee eee eee 3
2 Thalline and proper margins poorly developed;

ASCOSPOLES ZO—SOSCGHB UME 0.2 a ine ka ged G nore dG nore 24 Hare Ea Niel L. dilatatum
2 Thalline and proper margins well developed;

ASCOSPOKes OHI Gi 6S, ors ots diets ote Sew atenat aw annie andeutleer atendthte L. sericeum

3 Norstictic and stictic acids present; ascus 6-spored;

ascospores 7—12-locular, 35-45 x 10-15 um............. L. crassimarginatum
3 Stictic acid present; ascus 8-spored;

ascospores 7—9-locular, 25—40 x 8-10 um ................ L. melanostalazans

Acknowledgments

This research was supported by the National Natural Science Foundation of China
for the funds (No. 31270066 & 31093440) and Excellent Youthful Scientist Foundation
Project of Shandong Province (No. BS2011SW028). The authors are grateful to Dr.
Santosh Joshi (CSIR-National Botanical Research Institute, Lucknow, India) and Hua-
Jie Liu (College of Life Sciences, Hebei University, China) for their valuable comments
on the manuscript and for acting as presubmission reviewers.

Literature cited

Culberson CE 1972. Improved conditions and new data for the identification of lichen products by
a standardized thin-layer chromatographic method. Journal of Chromatography 72: 113-125.
http://dx.doi.org/10.1016/0021-9673(72)80013-X

Culberson CF, Kristensson H. 1970. A standardized method for the identification of lichen products.
Journal of Chromatography 46: 85-93. http://dx.doi.org/10.1016/S0021-9673(00)83967-9

Orange A, James PW, White FJ. 2001. Microchemical methods for the identification of lichens.
British Lichen Society, London, UK.

Staiger B. 2002. Die Flechtenfamilie Graphidaceae. Studien in Richtung einer natiirlicheren
Gliederung. Bibliotheca Lichenologica 85. 526 p.

Thrower SL. 1988. Hong Kong lichens. Urban Council, Hong Kong. 193 p.

Wei JC. 1991. An enumeration of lichens in China. International Academic Publishers, Beijing.
278 p.

MYCOTAXON

http://dx.doi.org/10.5248/130.253
Volume 130, pp. 253-258 January-March 2015

Periconiella liquidambaricola sp. nov. -
a new Chinese hyphomycete

UwE BRAUN”, STEFFEN BIEN, LyDIA HONIG, & BETTINA HEUCHERT

Martin-Luther- Universitat, Institut fiir Biologie, Bereich Geobotanik und Botanischer Garten,
06099 Halle (Saale), Germany

* CORRESPONDENCE TO: uwe.braun@botanik. uni-halle.de

ABSTRACT — The new species Periconiella liquidambaricola, collected on living leaves of
Liquidambar formosana (Altingiaceae) in Jiangxi Province, China, is described, illustrated
and compared with other species of the genus Periconiella. The new fungus is the first species
described from a host belonging to the Altingiaceae. Periconiella liquidambaricola is well
characterized by its very long conidiophores and smooth, pale scolecosporous conidia. It
has similar conidia to Periconiella lygodii on fronds of Lygodium flexuosum in India, but
differs in having conidia with attenuated to acute apexes and much longer conidiophores
with uninflated bases.

KEY worps — Ascomycota, asexual morph, Periconiella qualeicola, Asia

Introduction

Leaves of Liquidambar formosana infected by a range of plant pathogenic
micromycetes were recently collected during a biodiversity-ecosystem study
in a subtropical forest near Xingangshan, Jiangxi Province, China. One of
these fungi had conidiophores with repeatedly and densely branched apices,
pigmented conidiophores with several conspicuous conidiogenous loci, and
solitary, holoblastically formed conidia that was readily identifiable as a species
of the genus Periconiella Sacc. This fungus represents the first collection of a
Periconiella species on a host in the Altingiaceae. A comparison with other
species assigned to Periconiella revealed that the Chinese collections represent
an undescribed species.

Materials & methods

Light microscopy (OLyMPus BX50) was used to examine the material (unstained
samples mounted in distilled water, oil immersion, 1000x). The ranges of sizes for
conidiophores, conidia and other structures were based on 30 measurements. Digital

254 ... Braun & al.

images were made with a Ze1ss Axioskop 2 with ZE1ss AxioCam HR and occasionally
optimized with the software ZEIss AxioVision. Freehand drawings were made on the
basis of microscopic preparations.

Taxonomy

Periconiella liquidambaricola U. Braun, S. Bien & Honig, sp. nov. FIGS. 1, 2
MycoBank MB809832

Periconiellae lygodii morphologice similis sed maculis foliorum nullis, conidiophoris
multo longioribus, ad basim non inflatis et conidiis apice attenuato vel acuto.

Type: China, Jiangxi Province, Xingangshan, subtropical forest site of the BEF-China
Project, 29.1250° N, 117.9085° E, on living and wilting leaves of Liquidambar formosana
(Altingiaceae), 8 Sep. 2013, coll. S. Bien (HOLOTYPE-HAL 2673 F).

Erymo ocy: Derived from the name of the host genus (inhabitant of Liquidambar).

LEAF spots indistinct or lacking. CoLoNtgEs on living green leaves or on
diffuse, discolored patches of wilted leaves, yellow or ochraceous to pale brown.
MycELIvo internal and external; superficial hyphae emerge through stomata,
branched, straight to sinuous, 1-4 um wide, septate, subhyaline to medium
brown, thin-walled, verruculose. STROMATA not developed. CONIDIOPHORES
solitary, scattered, arise from superficial hyphae, at the top of mother cells,
erect, straight to slightly curved, 400-700 um long, unbranched stalk 5-10 um
wide, not swollen at the base, pluriseptate throughout, distance between septa
25-50 um, dark brown throughout or somewhat paler towards the apex, wall
thickened, 1-2 um, smooth, apical portion 2-5 times densely forked, branched
part 50-100 um long, primary branches about 25-70 um long, subcylindrical.
CONIDIOGENOUS CELLS integrated, terminal, 10-30 um long and 3-6 um wide,
subcylindrical or enlarged (subclavate), up to 9 um, straight to moderately
geniculate-sinuous, with a single or several conspicuous conidiogenous loci,

Fic. 1. Periconiella liquidambaricola. Micrographs of branched apical portions of
conidiophores: A. focussed on outline; B. focussed on conidiogenous loci; C. with adherent
and detached conidia. Bars: A, C = 50 um; B = 20 um.

Periconiella liquidambaricola sp. nov. (China) ... 255

BOE HESTON ESET REY CSESRIE

:
i
#
4
é
i
i
|

Fic. 2. Periconiella liquidambaricola.
A. superficial hyphae; B. solitary conidiophores arising from superficial hyphae;
C. branched apical parts of conidiophores; D. conidia. Bars: B, C. = 50 um; D = 10 um.

256 ... Braun & al.

circular, ring-like, 1.5-3 um diam, rim darkened-refractive, barely or slightly
thickened. Conrp1A solitary, obclavate, attenuated towards the tip, 50-140 x
4-7.5 um, 3-7-septate, sometimes constricted at the septa, pale brown, thin-
walled, smooth, apex subobtuse to pointed, base short obconically truncate,
hila 2-2.5 um wide, darkened-refractive, at least around the rim.

Discussion

Arzanlou et al. (2007) published a modern description of Periconiella and
revealed its phylogenetic afhnity within the Mycosphaerellaceae. Periconiella
velutina (G. Winter) Sacc., the type species, together with most other species
assigned to Periconiella are characterized by having amero-, didymo- or
phragmosporous conidia (Ellis 1967, 1971, 1976; Priest 1991; Hosagoudar &
Braun 1995; McKenzie 1996; Arzanlou et al. 2007; Mafia et al. 2008, etc.). All
known species of this genus are specialized and confined to hosts in particular
genera of plant families. Periconiella liquidambaricola is the first species
described from a host in the Altingiaceae.

The conidial shape places P. liquidambaricola in a small group of species
characterized by scolecosporous conidia. Periconiella lygodii Arch. Singh et
al. ex U. Braun (Braun 2004), described from India on Lygodium flexuosum,
is a morphologically similar species. A record of “Periconiella sp. close to
P. rapaneae M.B. Ellis” on Lygodium flexuosum from Myanmar (Thaung 2008)
seems to belong to P lygodii. Additional records of P. lygodii are known from
the Philippines (Begum et al. 2009) and Taiwan (Kirschner & Chen 2010),
in both cases on Lygodium japonicum, and from Laos on L. polystachyum
(Phengsintham et al. 2013). Periconiella lygodii is characterized by similar
smooth scolecosporous conidia, but forms distinct leaf spots, much shorter
conidiophores that are enlarged at the base, and conidia that are rounded at the
apex (Braun 2004).

Periconiella rapaneae M.B. Ellis on Rapanea in Ecuador (Ellis 1967, 1971) is
morphologically close to P. lygodii but differs in having long primary branches
(to 170 x 5-8 um). Periconiella araliacearum G.F. Laundon (Laundon 1972), P.
arcuata Arzanlou et al. (Arzanlou et al. 2007), P longispora Kamal et al. (Kamal
et al. 1979), P maianthemi R. Kirschner (Kirschner & Piepenbring 2008) and
P. santaloidis M.B. Ellis (Ellis 1967, 1971) are similar species. Periconiella
araliacearum differs in having much shorter conidiophores (to 200 um)
with sparingly branched apices, and long, verruculose conidia (to 170 um).
Periconiella longispora, described from India on Litsea chinensis, is characterized
by having long conidiophores (to 800 um) formed in fascicles, with branches up
to 175 um, and rather broad (36-180 x 4.5-9.5 um), smooth and 3-20-septate
conidia. Periconiella arcuata is distinct with its falcate, verruculose conidia,
and P. maianthemi is distinguishable by having much shorter conidiophores

Periconiella liquidambaricola sp. nov. (China) ... 257

(200-435 wm) and shorter verruculose conidia, (13.5-)19-43(-75) um.
Periconiella santaloidis differs in having conidiophores with densely branched
apices, short branchlets and verruculose conidia.

An additional comparable species is:

Periconiella qualeicola Dorn.-Silva & Dianese, nom. nov.
MycoBank MB809833
= Periconiella longispora Dorn.-Silva & Dianese, Mycologia 95: 1246, 2004 [“2003”],
nom. illeg.,
non P. longispora Kamal, Suj. Singh & R.P. Singh,1979.

This leaf-spotting hyphomycete described from Brazil (Dornelo-Silva &
Dianese 2004), can be distinguished by its much wider cylindrical conidia
(37-95 x 5-8 um).

Periconiella liquidambaricola seems to be rather common in the subtropical
forest site near Xingangshan, Jiangxi Province. Numerous leaf samples of
Liquidambar formosana were collected for mycological examination. Mycelium
with pigmented verruculose hyphae assignable to P liquidambaricola has been
found on most leaves, but well developed conidiophores have been observed on
a limited number of leaves.

Acknowledgments

We gratefully acknowledge funding by the German Research Foundation (DFG, FOR
891/1 and 891/2) and the National Science Foundation of China (NSFC, 30710103907
and 30930005). We are also much obliged to V.A. Mel'nik (Russia) and R.G. Shivas
(Australia) for pre-submission reviews.

Literature cited

Arzanlou M, Groenewald JZ, Gams W, Braun U, Crous PW. 2007. Phylogenetic and
morphotaxonomic revision of Ramichloridium and allied genera. Studies in Mycology 58: 57-
93. http://dx.doi.org/10.3114/sim2007.58.03

Begum MM, Shivas RG, Cumagun CJR. 2009. First record of Periconiella lygodii occurring on
Lygodium japonicum in the Philippines. Australasian Plant Pathology Notes 4: 17-18.

Braun U. 2004. Periconiella species occurring on ferns. Feddes Repertorium 115: 50-55.
http://dx.doi.org/10.2307/3761924

Dornelo-Silva D, Dianese JC. 2004. Hyphomycetes on the Vochysiaceae from the Brazilian cerrado.
Mycologia 95: 1239-1251. http://dx.doi.org/10.2307/3761924

Ellis MB. 1967: Dematiaceous hyphomycetes VIII. Mycological Papers 111: 1-46.

Ellis MB. 1971: Dematiaceous Hyphomycetes. Kew.

Ellis MB. 1976: More Dematiaceous Hyphomycetes. Kew.

Hosagoudar VB, Braun U. 1995. Two new Indian hyphomycetes. Indian Phytopathology 48:
260-262.

Kamal, Singh S, Singh RP. 1979. A new species of Periconiella from India. Current Science 48:
211-212.

Kirschner R, Chen CJ. 2010. Periconiella species (anamorphic Mycosphaerellaceae) from Taiwan.
Fungal Diversity 44: 135-148. http://dx.doi.org/10.1007/s13225-010-0054-0

258 ... Braun & al.

Kirschner R, Piepenbring M. 2008. Two new hyphomycetes from Panama. Mycological Progress 7:
21-29. http://dx.doi.org/10.1007/s11557-007-0549-6

Laundon GE. 1972. Records of fungal plant diseases in New Zealand - 2. New Zealand Journal of
Botany 9: 610-624. http://dx.doi.org/10.1080/0028825X.1971.10430224

Mafia RG, Ferreira EM, Ferreira FA, Braun U, Pereira OL. 2008. Periconiella lecythidis sp. nov.,
the causal agent of a zonate leaf spot disease of the Brazilian tree Lecythis pisonis. Mycological
Progress 7: 49-52. http://dx.doi.org/10.1006/s11557-007-0552-y

McKenzie EHC. 1996. A new species of Periconiella on Pomaderris. Mycotaxon 59: 43-45.

Phengsintham P, Chukeatirote E, McKenzie EHC, Hyde KD, Braun U. 2013. Monograph of
cercosporoid fungi from Laos. Current Research in Environmental and Applied Mycology 3:
24-158.

Priest MJ. 1991: Species of Periconiella and Stenella on Proteaceae in Eastern Australia. Mycological
Research 95: 924-927. http://dx.doi.org/10.1016/S0953-7562(09)

Thaung MM. 2008. A list of Hyphales (and Agonomycetes) in Burma. Australasian Mycologist 27:
149-172.

MY COTAXON

http://dx.doi.org/10.5248/130.259
Volume 130, pp. 259-264 January-March 2015

First record of Erysiphe syringae-japonicae in Turkey

ILGAZ AKATA* & VASYL P. HELUTA

‘Ankara University, Science Faculty, Department of Biology,
TR 06100, Ankara, Turkey

°M.G. Kholodny Institute of Botany, National Academy of Sciences of Ukraine,
2 Tereshchenkivska St., Kiev, 01601, Ukraine

‘CORRESPONDING AUTHOR: fungus@hotmail.com.tr

ABSTRACT — Erysiphe syringae-japonicae was reported on leaves of Syringa vulgaris for the
first time from Turkey. A short description, distribution, and illustrations for this powdery
mildew fungus are provided and discussed briefly.

Key worps — Asia Minor, Erysiphales, invasive species, lilac, Microsphaera

Introduction

A powdery mildew collected in Japan on the lilac, Syringa amurensis var.
japonica [= S. reticulata], was described by Braun (1982) as Microsphaera
syringae-japonicae (Erysiphales, Ascomycota). Later, this species was reported
from the Russian Far East (Bunkina 1991, as “Microsphaera syringae”) and from
Korea (Shin 2000). Microsphaera syringae-japonicae was already known on
lilacs in North America and Europe, and was distinguished from M. syringae,
mainly by its evanescent mycelium, its larger number of spores in the ascus,
and its usually more extensively pigmented chasmothecial appendage bases.

In 1988, one of the authors (VP Heluta) critically examined the type specimens
of powdery mildews described from the Russian Far East, and showed that
one of them, the type specimen of M. aceris on leaves of Acer barbinerve, had
chasmothecia identical to those of M. syringae-japonicae. However they were in
adherent groups that could have drifted from another host such as a Syringa sp.
Thus, M. aceris is an earlier synonym for M. syringae-japonicae and has priority
within the genus Microsphaera Lév. However, as already discussed by Braun et
al. (2003) and Braun & Cook (2012), Microsphaera is now considered a section
of the genus Erysiphe DC.; as the epithet aceris is already occupied in Erysiphe,
the correct current name is E. syringae-japonicae |= M. syringae-japonicae].

260 ... Akata & Heluta

In the 1990s, E. syringae-japonicae from East Asia was introduced into
Europe (Seko et al. 2008, 2011), where it is now common in Germany, Poland,
Russia, Ukraine, Switzerland, etc. (Braun 1998, 2002, Piatek 2003, Heluta 2006,
Rusanov & Bulgakov 2008, Braun & Cook 2012). However, it had not been
reported in either the European or Asian parts of Turkey until autumn 2012,
when one of the authors (I. Akata) collected it on Syringa vulgaris in Ankara.

Materials & methods

Chasmothecia were studied on herbarium material dried between papers at 22-24°C.
They were examined in distilled water and photographed under a light microscope
«Primo Star» (Carl Zeiss, Germany) using the Camera «Canon A 300» and software
«AxioVision 4.7». Digital data were analysed statistically with n 230 for each character.
For study under the scanning electron microscope (JSM-6060LA, Japan), dried pieces
of infected Syringa leaves were glued onto the metal stubs and coated with gold.

Identification was made using the descriptions of Braun & Cook (2012). In addition,
the specimen was compared with samples of E. syringae-japonicae collected in Germany,
Switzerland, Ukraine, and the Russian Far East. The studied materials are deposited in
the National Herbarium of M.G. Kholodny Institute of Botany, National Academy of
Sciences of Ukraine, Kiev, Ukraine (KW).

Taxonomy

Erysiphe syringae-japonicae (U. Braun) U. Braun & S. Takam., Schlechtendalia 4:
14. 2000. Figs 1, 2
= Microsphaera syringae-japonicae U. Braun, Mycotaxon 15: 121. 1982.
= Microsphaera aceris Bunkina, Komarovskie Chteniya 21:
82. 1974 [non Erysiphe aceris DC. 1815].
= Erysiphe acerina U. Braun & S. Takam., Schlechtendalia 4: 5. 2000.

Mycelium amphigenous, white, often effused or in patches, evanescent
to sub-persistent. Hyphae thin, up to 7 um wide. Hyphal appressoria well
developed, lobed, in opposite pairs or sometimes single. Chasmothecia brown
to dark brown, scattered to gregarious on the leaf surface, 70-90 um diam.
Peridium cells polygonal to almost rounded, 10-16 x 18-21 um. Appendages
4-12, equatorial, aseptate, stiff, usually straight to rarely curved, smooth
to rough, thick-walled, hyaline but brownish or brown at the base, 75-130
um long, apices 4-5 times regularly dichotomously branched often in three
dimensions (ie., branched part not fully flat), rather compact, sometimes
primary and secondary branches elongated and therefore apical parts somewhat
deeply forked, tips recurved. Asci 3-6, sessile or short-stalked, broadly ellipsoid
or slightly ovoid, hyaline, 40-55 x 30-40 um, (5-) 6-8-spored. Ascospores
hyaline, ellipsoid or ovoid, 16-22 x 11-14 um.

SPECIMEN EXAMINED — TURKEY, ANKARA, Tandogan, 39°56’N 32°49’E, alt. 860 m, on
Syringa vulgaris L. (Oleaceae), 17.10.2012, Ilgaz Akata (KW 40328F).

Erysiphe syringae-japonicae new for Turkey ... 261

\ A 2 i .
DOM w |
ef Xk rae Ne yf Me
Wer oN i “~M ]
RON Bt S
anamennemr Sy , |
y Sehr 4 %
y i f
Wy
., oe ry

4 > . a . on. \ eS ¥ —— SSS
pschaLy Spin = Ve 36k URS 21 S88 41 Grim
/ { S 5 , oe

Fic. 1. Erysiphe syringae-japonicae (KW 40328F) on Syringa vulgaris: a - chasmothecia on the host
plant leaf (upper surface, in reflected light); b, c - chasmothecia splitting with release of ascospores
(b) and asci (c) and with appendages colored at the base; d, e - chasmothecia with appendages
(SEM); f - surface of the peridium (SEM).

Discussion

Many Syringa species are cultivated in different countries of the northern
hemisphere as important ornamental plants. Some of them are susceptible to
powdery mildew caused by Phyllactinia fraxini (DC.) Fuss and two species of
Erysiphe sect. Microsphaera. Phyllactinia fraxini is uncommon and causes little
harm to its hosts. The other two species, E. syringae and E. syringae-japonicae,

262 ... Akata & Heluta

a
i> 488 164m ¢

- a -

Fic. 2. Erysiphe syringae-japonicae (KW 40328F): g - basal part of chasmothecial appendage
(SEM); h, i - apical parts of appendages (SEM); j - paired hyphal appressoria (SEM); k - asci;
1 - ascospores.

are common, and E. syringae-japonicae especially can harm the hosts. Seko et
al. (2008, 2011) studied the worldwide spread of these fungi and found the
origin of E. syringae to be North America from whence it was introduced to
Europe, East Asia, and South America. However, recently the fungus has been
supplanted by the Asian E. syringae-japonicae, particularly in Europe. The last
time E. syringae was collected in Ukraine was in 2003 (Seko et al. 2011), where
it formed only the anamorph.

Erysiphe syringae-japonicae new for Turkey ... 263

Erysiphe syringae and E. syringae-japonicae are different genetically (only
94% similarity between the ITS sequences of the two species; see Seko et al.
2008) as well as morphologically. Erysiphe syringae has typically 4-5-spored
asci and nearly colorless appendages, while the asci of E. syringae-japonicae
contain (5-)6-8 spores and its appendages are more or less pigmented from
their base up to the middle part. Thus, these species can be easily distinguished
by morphological analysis.

Lilac (Syringa vulgaris) is also cultivated in Turkey as an ornamental plant.
According to the recent literature on Turkish powdery mildews in Turkey,
only E. syringae has been previously reported on this plant (Bahcecioglu &
Yildiz 2005; Bahcecioglu et al. 2006; Kabaktepe & Bahcecioglu 2006, 2009;
Kavak 2011; Severoglu & Ozyigit 2012), and there is no record of E. syringae-
japonicae. However, our sample is identical to the specimens collected in
Europe, especially in Ukraine. Thus, the spread of E. syringae-japonicae in
Turkey needs more research and monitoring.

Acknowledgments

The authors are grateful to Uwe Braun (Germany) for kindly sending several
specimens of E. syringae-japonicae collected in West Europe and for his pre-submission
review. We thank VI Sapsay (Ukraine) for his help with scanning electron microscopy.
We also thank Uwe Braun, Roger Cook, Wieslaw Mulenko, and Shaun Pennycook for
their helpful comments and careful review of this article.

Literature cited

Braun U. 1982. Descriptions of new species and combinations in Microsphaera and Erysiphe (II).
Mycotaxon 15: 121-137.

Braun U. 1998. Neufunde echter Mehltaupilze (Erysiphales) aus der BR Deutschland.
Schlechtendalia 1: 31-40.

Braun U. 2002. Erysiphe miurae and E. syringae-japonicae — new records from Russia. Mikologiya
i Fitopatologiya 36(2): 15-16.

Braun U, Cook RTA. 2012. Taxonomic manual of the Erysiphales (powdery mildews). CBS
Biodiversity Series 11. 707 p.

Braun U, Cunnington JH, Brielmaier-Liebetanz U, Ale-Agha N, Heluta V. 2003. Miscellaneous
notes on some powdery mildew fungi. Schlechtendalia 10: 91-95.

Bahcecioglu Z, Yildiz B. 2005. A study on the microfungi of Sivas Province. Turkish Journal of
Botany 29: 23-44.

Bahcecioglu Z, Kabaktepe S, Yildiz B. 2006. Microfungi isolated from plants in Kahramanmaras
Province, Turkey. Turkish Journal of Botany 30: 419-434.

Bunkina IA. 1991. Poriadok Erysiphales. Nizshie rastenia, griby i mokhoobraznyie sovetskogo
Dal’nego Vostoka. Griby, Vol. 2. Askomitsety. Erizifalnyie, klavitsipital’nyie, gelotsial’nyie.
Nauka (Leningrad): 11-142.

Heluta VP. 2006. Boroshnystorosiani hryby (poriadok Erysiphales) Kanivs’koho pryrodnoho
zapovidnyka. Zapovidna sprava v Ukraini 12(2): 23-32.

264 ... Akata & Heluta

Kabaktepe S, Bahcecioglu Z. 2006. Microfungi identified from the flora of Ordu Province in Turkey.
Turkish Journal of Botany 30: 251-265.

Kavak H. 2011. Two new records of powdery mildews with their effectiveness on three ornamentals
in Turkey. African Journal of Agricultural Research 6(5): 1076-1079.
http://dx.doi.org/10.5897/AJAR10.992.

Piatek M. 2003. Erysiphe azaleae and Erysiphe syringae-japonicae introduced in Poland. Mycotaxon
87: 121-126.

Rusanov VA, Bulgakov TS. 2008. Muchnistorosianye griby Rostovskoi oblasti. Mikologiya i
Fitopatologiya 42(4): 314-322.

Seko Y, Bolay A, Kiss L, Heluta V, Grigaliunaite B, Takamatsu S. 2008. Molecular evidence in
support of recent migration of a powdery mildew fungus on Syringa spp. into Europe from
East Asia. Plant Pathology 57(2): 243-250. http://dx.doi.org/10.1111/j.1365-3059.2007.01775x.

Seko Y, Heluta V, Grigaliunaite B, Takamatsu S. 2011. Morphological and molecular characterization
of two ITS groups of Erysiphe (Erysiphales) occurring on Syringa and Ligustrum (Oleaceae).
Mycoscience 53(3): 174-182. http://dx.doi.org/10.1007/s10267-010-0088-x.

Severoglu Z, Ozyigit I. 2012. Powdery mildew disease in some natural and exotic plants of Istanbul,
Turkey. Pakistan Journal of Botany 44: 387-393.

Shin H.-D. 2000. Erysiphaceae of Korea. National Institute of Agricultural Science and Technology,
Suwon. 321 p.

MY COTAXON

http://dx.doi.org/10.5248/130.265
Volume 130, pp. 265-273 January-March 2015

AFLP characterization of three argentine Coprotus species

ARACELI MARCELA RAMOS’, LUIS FRANCO TADIC', NAHUEL POLICELLI’,
LAURA INES FERREYRA?’, & ISABEL ESTHER CINTO’

' Lab 9 (PROPLME-PRHIDEB-CONICET), Departamento de Biodiversidad y Biologia
Experimental, Facultad de Ciencias Exactas y Naturales,
Universidad de Buenos Aires, Ciudad Universitaria, 1428, Buenos Aires, Argentina

? Departamento de Ecologia, Genética y Evolucion. Facultad de Ciencias Exactas y Naturales,
Universidad de Buenos Aires, Ciudad Universitaria, 1428, Buenos Aires, Argentina

* CORRESPONDENCE TO: icinto@bg.fcen.uba.ar

AxsstRact — AFLP methodology was applied to characterize three Coprotus species
(C. lacteus, C. niveus, C. sexdecimsporus) so as to estimate the levels of polymorphism within
species, to analyze the phenetic relationships among them, and to contrast the AFLP findings
to those of a previous RAPD study. The high number of AFLP bands obtained with the six
assayed primers allowed us to detect intra-specific variability. The genetic variability within
species obtained using AFLP (measured in terms of percentage of polymorphic loci) was two
to three times higher than those obtained by RAPD. The phenograms generated by AFLP
markers grouped all strains of the same species into three defined clusters, and a higher
association between C. lacteus and C. sexdecimsporus was also observed. The AFLP technique
could become a powerful tool for genera such as Coprotus, in which a high intra-specific
homogeneity does not allow detection of genetic variability using other PCR-based markers.

KEY worps — Ascomycota, molecular markers, coprophilous

Introduction

The fungal genus Coprotus Korf & Kimbr. comprises coprophilous
discomycetes, which belong to the Pyronemataceae Corda. Species of this
genus are homothallic, probably due to an adaptation to the substrate, which
determines their sexual isolation (Wicklow 1981). Sexual reproduction
produces ascospores dispersed by effective discharge methods.

A homothallic mating system can be very difficult to distinguish from
clonality using some molecular markers, since both systems produce progenies
that do not show evidence of segregation for these markers and yield linkage
disequilibrium at the population level over the long term (Billiard et al. 2012).

266 ... Ramos & al.

Coprotus species have been traditionally distinguished by such cytological
and morphological characters (Kimbrough et al. 1972) as the number of
ascospores per ascus, the presence or absence of pigments in paraphyses and
excipulum, and the size and shape of asci, ascospores and sterile elements.

Different concepts have been used to define fungal species. The phenotypic
concept is the classic approach based on morphology, and Coprotus species
have traditionally been classified on cytological and morphological characters
(Kimbrough et al. 1972). However, difficulties often arise while attempting to
identify Coprotus species, as they are morphologically very similar, characters
frequently overlap, and some vary greatly according to the culture conditions
(Dokmetzian et al. 2005). A polythetic concept, which circumscribes species
based on a combination of characters (Kohn 1992, Guarro et al. 1999), could
prove helpful in characterizing the Coprotus genus.

RAPD [random amplified polymorphic DNA] analysis was performed on
Coprotus species by Ramos et al. (2008). In that study, the RAPD technique
confirmed the previous identification of strains using morphological
characters (Dokmetzian et al. 2005) and provided a greater number of species-
specific bands, but genetic variability within species (in terms of percentage
of polymorphic loci) was very low. All strains had the same geographic origin
and showed identical patterns for the six primers assayed on the three Coprotus
species studied. Therefore, it would seem that to detect variability among
strains, other PCR markers, such as AFLP, should be used.

The amplified fragment length polymorphism (AFLP) method (Vos
et al. 1995) is a DNA-based fingerprinting technique, which requires no
prior sequence information on the organism under scrutiny. This method is
used to determine the degree of similarity among isolates and offers certain
advantages over other techniques, i.e., high level of identified polymorphism,
high reproducibility, and relative technical simplicity (Briad et al. 2000). AFLP
analysis has been successfully used to estimate the amount of genetic variation
within fungal species. Given that a large number of amplicons can be screened,
AFLP is especially useful for characterizing clonal lineages and for establishing
phenetic relationships among species with little morphological differentiation.
AFLP has a clear advantage over other PCR based markers such as RAPDs, not
only in terms of reproducibility but also because more loci can be screened in
each reaction.

In our present study we applied the AFLP methodology to characterize three
Coprotus species—C. lacteus (Cooke & W. Phillips) Kimbr., C. niveus (Fuckel)
Kimbr., and C. sexdecimsporus (P. Crouan & H. Crouan) Kimbr. & Korf—so as
to estimate the levels of polymorphism within species, to analyze the phenetic
relationships among them, and to contrast the findings of our AFLP study to
those of a previous RAPD study.

Coprotus AFLP characters [Argentina] ... 267

TABLE 1. Fungal strains of Coprotus species used, with their Argentinian collection
localities and BAFC numbers

STRAIN GEOGRAPHICAL LOCATION * BAFC
C. lacteus
lacA1(11), lacA3(12), lacA6(13), lacA10(14), Agronomia, CABA 874, 1937, 1940,
lacA13(15) 1941, 1942
lacL1(16), lacL 3 17), lacL4(18), lacL6 (19) Villa Lugano, CABA 1944, 1945, 1946,
1947
C. niveus
nivBC2 (25), nivBC3 (27), nivBC4 (30) Bahia Craft, Villa La Angostura, NQ 1970, 1971, 1972
nivE1 (48), nivE2 (49) Bahia Ensenada, TF 1956, 982
nivC2 (42), nivC3 (38), nivC4 (45), Campana, PBA 1960, 1961, 1962,
nivC5 (46) 1963
nivU1 (36), nivU3 (37), nivU6 (39), Ciudad Universitaria, CABA 1964, 1965, 1966,
nivU7 (40), nivU8 (41) 1967, 1968
nivL1 (34), nivL3 (35) Villa Lugano, CABA 1973, 1974
C. sexdecimsporus
sexU 1 (6), sexU2 (7), sexU4 (8) Ciudad Universitaria, CABA 1952, 1953, 1954
sexG1 (5), sexG4 (21), sexG7 (23) Los Gigantes, C 1948, 1950, 1951

* C = Cordoba province:, CABA = Buenos Aires city; NQ = Neuqueén province;
PBA = Buenos Aires province; TF = Tierra del Fuego province.

Materials & methods

Isolation and maintenance of monosporic strains

The characters used to identify the Coprotus species were those used by Kimbrough et
al. (1972). The isolation and maintenance of monosporic strains followed the procedure
indicated by Suarez et al. (2006). Thirty-one monosporic strains, from nine geographical
locations, were used (TABLE 1). All strains were deposited in the Herbarium and Culture
Collection of the Department of Biodiversity, Faculty of Natural & Exact Sciences,
University of Buenos Aires, Argentina (BAFC).

DNA extraction

Mycelium was obtained as in Ramos et al. (2000) and ground to powder in liquid
nitrogen using a sterile pestle. Genomic DNA was extracted following Gottlieb &
Lichtwardt (2001). Quality control and quantification of genomic DNA was carried out
by agarose gel (0.8% w/v) electrophoresis and by comparison with a DNA molecular-
size standard (Lambda EcoRI/HinDII, Promega Corp.). Ethidium bromide gels were
photographed under UV light.

AFLP

The AFLP methodology was carried out on 250 ng of genomic DNA using the
AFLP® Analysis System for Microorganisms Primer Kit (Invitrogen) as described in the
instructions manual with minor modifications (Gottlieb et al. 2005). Selective primers
were combined as in TABLE 2.

268 ... Ramos & al.

TABLE 2. AFLP primers used in this study

PRIMERS SEQUENCE 5’-3’

E+ACG GAC TGC GTA CCA ATT CAC G
E+AAC GAC TGC GTA CCA ATT CAA C
E+AAG GAC TGC GTA CCA ATT CAA G
M+A GAT GAG TCC TGA GTA AA
M+G GAT GAG TCC TGA GTA AG
M+C GAT GAG TCC TGA GTA AC
M+T GAT GAG TCC TGA GTA AT

* E stands for EcoRI and M stands for Msel.

All PCR amplifications were performed in a TECHNE PROGENE thermal cycler.
Polyacrylamide gel electrophoresis conditions followed Gottlieb et al. (2005). A
30-330 bp AFLP® DNA Ladder (Invitrogen) size marker was included twice in each
electrophoresis, and the size of AFLP bands scored ranged from 90 to 330 bp. AFLP
bands were visualized using the SILVER SEQUENCE TM DNA Sequencing System
(Promega).

Statistical methods

The data were extracted as a table and marked as either present (1) or absent (0).
Monomorphic bands (bands present in all individuals of a species) were discriminated
within each species and across the entire data set. The binary matrix was analyzed
with the program NTSYS-PC version 2.02 (Rohlf 1993). The unweighted pair-group
arithmetic mean method (UPGMA) cluster analysis was performed on the simple
matching (SM) association coefhicient (Sneath & Sokal 1973), and the same program
generated the phenogram showing similarity relations. The distortion produced during
the grouping analysis was calculated using the correlation cophenetic coefficients
(r) (Sokal & Rohlf 1962) using the NTSYSPC version 2.02 (Rohlf 1993) program. A
cophenetic correlation (r) of 0.8 was considered a good fit. A three-dimensional graphic
was obtained with the principal coordinated ordination method (Gower 1966).

The percentage of polymorphic loci (P = (number of polymorphic loci/ number of
loci analyzed) x 100) for each primer combination was calculated.

Results

The analysis of the six primers that produced clear and reproducible bands
by AFLP amplification yielded a total of 926 bands that appeared consistently
in all experiments among the amplified fragments of the 31 isolates. Although
the amplified bands ranged from 50 to 550 bp, we analyzed only bands from
90 to 330 bp.

TABLE 3 describes the total number of AFLP (polymorphic + monomorphic)
bands and the percentage of polymorphic loci obtained for each selective
primer combination in the three assayed fungal species. All primers detected
polymorphic bands within species and a high percentage of polymorphic loci

Coprotus AFLP characters [Argentina] ... 269

TABLE 3. Total number of AFLP (polymorphic + monomorphic) bands detected for
each selective primer combination per Coprotus species assayed.

PRIMER SPECIES ToTaL N° oF POLYMORPHIC MONOMORPHIC P%
COMBINATION BANDS BANDS BANDS
M+T/E+ACG C. lacteus 50 16 34 32
C. niveus 27 7 20 26
C. sexdecimsporus 72 60 12 83
M+C/E+AAG C. lacteus 101 28 73 28
C. niveus 108 10 98 9
C. sexdecimsporus 163 141 22 86
M+A/E+AAG C. lacteus 75 12 63 16
C. niveus 90 38 52 42
C. sexdecimsporus 152 135 17 89
M+C/E+ACG C. lacteus 77 23 54 30
C. niveus 129 75 54 58
C. sexdecimsporus 156 144 12 92
M+G/E+ACG C. lacteus 79 26 53 33.
C. niveus 80 36 44 45
C. sexdecimsporus 100 74 26 74
M+A/E+ACG C. lacteus 74 24 50 32
C. niveus 70 14 56 20
C. sexdecimsporus 143 130 13 90

were obtained for C. sexdecimsporus and a reasonable percentage for C. niveus
and for C. lacteus.

Genetic variability within species, measured in terms of percentage of
polymorphic loci was variable: P = 87 % for C. sexdecimsporus, P= 36 % for C. niveus
and, P= 28 % for C. lacteus.

The primer group M+C/E+ACG was more effective in detecting
polymorphisms (TABLE 3).

Degree of similarity and cluster analysis

The phenogram obtained using the UPGMA method was constructed
based on Simple Matching (SM) coefficients. Little distortion occurred while
constructing this phenogram, as implied by the value of the correlation
cophenetic coefficient (r=0.991). There was no direct correlation between
molecular genotype and geographic origin.

The UPGMA phenogram (Fic. 1) grouped all strains in three defined
clusters that correspond to the three Coprotus species.

The degree of similarity among species was low. Between C. lacteus and
C. sexdecimsporus, the coefficient of similarity was S = 0.5. The group formed

270 ... Ramos & al.

C5 rls

SG3

sna}dbd] ‘D>

wn
(a)
A
VBA!
a
snuodsw2apxas 9)

SG1

8
snanlu ‘>

0.73
Coefficient

FiGurE 1. UPGMA phenogram showing relationships among Coprotus lacteus, C. sexdecimsporus,
and C. niveus isolates based on the simple matching (SM) association coefficient estimated from
AFLP loci.

by the isolates identified as C. niveus clustered with the other two species with
a coefficient of similarity of S = 0.46.

The main group (Fic. 1, group Gl) comprised two subgroups (SG1 and
SG2) formed by all isolates of C. niveus. Subgroup SG1 is formed by two sets
(Cl and C2) and SG2 subgroup comprised C3 and C4 sets and one isolate
separated from the rest. A second group (Fic. 1, G2) included two subgroups
(SG3 and SG4). Subgroup SG3 is formed by two sets (C5 and C6) that included
all isolates of C. lacteus. The other subgroup (SG4) comprised C7 and C8 sets,
formed by all isolates of C. sexdecimsporus.

Intra-specific variability detected for C. lacteus and C. niveus was low. This
fact is reflected by the association coefficient: SM = 0.91 among isolates of
C. lacteus and SM = 0.92 among C. niveus isolates. Only two pairs of isolates
presented 100% of similarity (association coefficient 1.0), and they belonged to
C. lacteus. Coprotus sexdecimsporus showed a higher degree of variability than
the other species, with SM = 0.54 among all isolates of this species.

The ordination of isolates through the principal coordinated method
allowed us to recognize three groups (Fic. 2, groups 1-3) in three-dimensional
dispersion revealing nearly the same relations between isolates as the
phenogram (Fic. 1). The first group included all isolates of C. lacteus, very
closely attached in the three axes. The second set showed C. sexdecimsporus
isolates differentiated in axis 1 but very closely in the other two axes. The third
comprised isolates of C. niveus closely together in the three axes, thus revealing
a high degree of similarity.

Coprotus AFLP characters [Argentina] ... 271

G Il

C. sexdecimsporus

GIll GI

C. niveus

C. lacteus

FiGuRE 2. Three-dimensional graphic of Coprotus species obtained with the Principal Coordinates
ordination technique.

Discussion

In the present study the high number of AFLP bands obtained with the six
primers assayed allowed us to detect intra-specific variability.

Previous biochemical and RAPD studies detected a high intra-specific
homogeneity (Suarez et al. 2006, Ramos et al. 2008), which could be related
to the fact that Coprotus is homothallic and produces clonal offspring (Rayner
1994). In the present study the genetic variability within species, in terms of
percentage of polymorphic loci, was two to three times higher than those
obtained by RAPD. AFLP fingerprinting is more effective for detecting genetic
diversity in species with high intra-specific homogeneity (e.g., Coprotus),
because it generated a very large number of polymorphic loci.

Genetic diversity could be a potential indicator of the relative abundance of
sexual and asexual reproduction (Chen & McDonald 1996). Furthermore other
alternatives for genetic interaction among fungi besides sexual recombination
are heterokaryosis and mitotic recombination (Esser & Kuenen 1967). In this

272 ...Ramos & al.

sense, natural selection could lead to genetic divergence and therefore increase
genetic variability levels in the homothallic fungi as Coprotus (non-out crossing
populations). The AFLP technique allowed us to detect such intraspecific
variability.

In the phenograms generated by AFLP and RAPD markers, all strains of
the same species group into three defined clusters. Besides, both phenograms
showed very similar phenetic relationships among species.

The phenogram obtained from AFLP datasets showed a higher association
between C. lacteus and C. sexdecimsporus. This result is consistent with that
obtained by means RAPD markers. This agreement might be related to the
nature of these markers since AFLP and RAPD are usually considered neutral
markers. Sampling and genotyping the progeny within sexual structures in
natural populations should allow determining whether haploid self-mating
system actually occurs in nature (Billiard et al. 2012).

The AFLP technique could become a powerful tool for researching genera
with homothallic mating systems such as Coprotus, in which a high intra-
specific homogeneity does not allow detection of variability using other PCR-
based markers.

Acknowledgements

This study was supported by a grant from Consejo Nacional de Investigaciones
Cientificas y Técnicas (CONICET) and University of Buenos Aires, Argentina. The
authors thank Dr. Lina Bettucci (Universidad de la Republica, Montevideo, Uruguay)
and Dr. Alejandra Fazio (Universidade de Sao Paulo, Brasil) for expert presubmission
review.

Literature cited

Billiard $, Lépez-Villavicencio M, Hood ME, Giraud T. 2012. Sex, outcrossing and mating types:
unsolved questions in fungi and beyond. Journal of Evolutionary Biology 25: 1020-1038.
http://dx.doi.org/10.1111/j.1420-9101.2012.02495.x

Briard M, Le Clerc V, Grzebelus LD, Senalik D, Simon PW. 2000. Modified protocols for rapid
carrot genomic DNA extraction and AFLP ™ analysis using silver stain or radioisotopes. Plant
Molecular Biology Reporter 18: 235-241. http://dx.doi.org/10.1007/BF02823994

Chen RS, McDonald BA. 1996. Sexual reproduction plays a major role in the genetic structure of
populations of the fungus Mycosphaerella graminicola. Genetics 142: 1119-1126.

Dokmetzian DA, Ramos AM, Cinto IE, Suarez ME, Ranalli ME. 2005. Seis especies del género
Coprotus (Pyronemataceae) de Argentina estudiadas en cultivo. Hickenia 3(57): 243-252.

Esser K, Kuenen R, 1967. Genetics of Fungi. Springer-Verlag, New York.
http://dx.doi.org/10.1007/978-3-642-86814-6

Gottlieb AM, Lichtwardt RW. 2001. Molecular variation within and among species of Harpellales.
Mycologia 93: 65-80. http://dx.doi.org/10.2307/3761606

Gottlieb AM, Giberti GC, Poggio L. 2005. Molecular analyses of the genus Ilex (Aquifoliaceae) in
southern South America, evidence from AFLP and ITS sequence data. American Journal of
Botany 92: 352-369. http://dx.doi.org/10.3732/ajb.92.2.352

Coprotus AFLP characters [Argentina] ... 273

Gower JC. 1966. Some distance properties of latent root and vector methods used in multivariate
analysis. Biometrika 53: 325-338. http://dx.doi.org/10.1093/biomet/53.3-4.325

Guarro J, Gené J, Stchigel AM. 1999. Developments in fungal taxonomy. Clinical Microbiology
Reviews 12: 454-500.

Kimbrough JW, Luck-Allen ER, Cain RE. 1972. North American species of Coprotus (Thelebolaceae:
Pezizales). Can. J. Bot. 50: 957-971. http://dx.doi.org/10.1139/b72-116

Kohn LM. 1992. Developing new characters for fungal systematics: an experimental approach for
determining the rank of resolution. Mycologia 84: 139-153. http://dx.doi.org/10.2307/3760244

Majer D, Mithen R, Lewis BG, Vos P, Oliver RP. 1996. The use of AFLP fingerprinting for the
detection of genetic variation in fungi. Mycol. Res. 100: 1107-1111.
http://dx.doi.org/10.1016/S0953-7562(96)80222-X

McDonald BA, Linde C. 2002. Pathogen population genetics, evolutionary potential, and
durable resistance. Ann. Rev. Phytopathol. 40: 349-379. http://dx.doi.org/10.1146/annurev.
phyto.40.120501.101443

Ramos AM, Forchiassin F, Ranalli ME, Saidman B. 2000. Isozyme analysis of different species of the
genus Saccobolus (Ascomycetes, Pezizales). Mycotaxon, 74: 447-462

Ramos. AM, Dokmetzian DA, Ferreyra LI, Ranalli ME. 2008. First report on RAPD patterns able
to differentiate some Argentinian species of the genus Coprotus (Ascomycota). Mycotaxon, 103:
9-19.

Rayner AD. 1994. Evolutionary processes affecting adaptation to saprotrophic life styles in
ascomycete populations. 261-271, in: DL Hawksworth (ed.). Ascomycete Systematics.
Problems and Perspectives in the Nineties. NATO ASI series. Plenum Press, USA.

Rohlf JE. 1993. NTSYS-PC. Numerical taxonomy and multivariate analysis system. Version 1.8.
New York: Exeter Software Applied Biostatistics Inc.

Sneath PH, Sokal RR. 1973. Numerical taxonomy: the principles and practice of numerical
classification. San Francisco: W.H. Freeman & Co.

Suarez ME, Ranalli ME, Dokmetzian DA, Ramos AM. 2006. Characterization of three species of
the genus Coprotus (Ascomycota) by isozyme analysis. Mycotaxon 97: 257-273

Sokal RR, Rohlf JF. 1962. The comparison of dendrograms by objective methods. Taxon 11: 33-40.
http://dx.doi.org/10.2307/1217208

Vos P, Hogers R, Bleeker M, Reijans M, Lee TV, Hornes M, Friters A, Pot J, Paleman J, Kuiper
M, Zabeau M. 1995 AFLP: a new technique for DNA fingerprinting. Nucl. Acids Res. 23:
4407-4414. http://dx.doi.org/10.1093/nar/23.21.4407

Wicklow DT. 1981 The coprophilous fungal community: a mycological system for examining
ecological ideas. 47-76, in: DT Wicklow, GC Carroll (eds). The Fungal Community: its
Organization and Role in the Ecosystem. Marcel Dekker, USA.

MY COTAXON

http://dx.doi.org/10.5248/130.275
Volume 130, pp. 275-278 January-March 2015

Verticicladus hainanensis, a new aquatic hyphomycete

MING-TIAN GuoS, MIN QIAOS, JIAN- YING Li, WEI WANG, & ZE-FEN YU*

Laboratory for Conservation and Utilization of Bio-resources,
Key Laboratory for Microbial Resources of the Ministry of Education,
Yunnan University, Kunming, Yunnan, 650091, P. R. China

*CORRESPONDENCE TO: Zfyuqm@hotmail.com

ABSTRACT —Verticicladus hainanensis, isolated from decaying leaves in Jianfengling Nature
Reserve, Hainan Province, China, is described and illustrated. It is characterized by fusoid,
5-8-septate, slightly grey to dark fuliginous conidia.

Key worps —fungal diversity, morphology, freshwater fungi, taxonomy

Introduction

The fungal genus Verticicladus is characterized by rhexolytic conidial
liberation from conidiogenous cells verticillately fasciculated on short
cylindrical conidiophores (Matsushima 1993). Two species have been
described, V. amazonensis Matsush. and V. subiculifer Matsush. (Matsushima
(99351996),

China has an enormous fungal diversity occurring on dead branches, rotten
submerged wood, and leaves. Many anamorphic fungi collected in southern
China have been published recently (Zhang et al. 2009a,b,c, 2011, 2012; Ma et
al. 2010, 2011a,b, 2012a,b; Wang et al. 2011; Ren et al. 2011, 2012). A species
isolated from decaying leaves in a small river from Jianfengling Nature Reserve,
Hainan Province, China had the characteristics of the genus Verticicladus but
differed from the two described species. Here, the new species is described and
illustrated in detail.

Materials & methods

The culture was isolated from dicotyledonous leaves submerged in a river in
Hainan. The rotten leaves were scissored to several 2-4 x 2-4 cm sized fragments and
then spread on to the surface of CMA (20 g cornmeal, 18 g agar, 40 mg streptomycin,

‘Min Qiao & Ming-Tian Guo contributed equally to this work

276 ... Guo, Qiao, & al.

30 mg ampicillin, 1000 ml distilled water) medium for ten days; single conidia were
isolated using sterilized needles while viewing with a BX51 microscope, and cultivated
on CMA in Petri plates. Morphological observations were made from CMA after
incubation at 28°C for one week; pure cultures and a permanent slide were deposited
in the Herbarium of the Laboratory for Conservation and Utilization of Bio-resources,
Yunnan University, Kunming, Yunnan, P.R. China (YMF; formerly Key Laboratory of
Industrial Microbiology and Fermentation Technology of Yunnan).

Taxonomy

Verticicladus hainanensis M.T. Guo & Z.F. Yu, sp. nov. PLATE 1
MycoBank MB 809830
Differs from Verticicladus amazonensis by its shorter, fusiform conidia and from V.

subiculifer by its attenuate conidia.

Type: PR China, Hainan Province, Jianfengling Nature Reserve, Chinese Academy of
Science, 18°44’N 108°49E, elev. 682 m, in a river on submerged leaves of an unidentified
dicotyledonous plant, 23 June 2011, G. Z. Yang (holotype, YMF1.04024 [dried agar
culture]; permanent slide, YMF1.040241; ex-type cultures, YMF1.040242).

EryMo toy: hainanensis refers to the province in which the species was found.

COLONIES growing moderately on CMA, attaining about 15 mm diam. after
7 days cultivation on CMA at 28°C. Colony effuse, dark olive to fuliginous,
mycelium partly superficial, partly immersed in substratum, composed of
branched, septate, smooth hyphae. Vegetative hyphae 1.9-3.8 um wide, often
2.5-3.5 um wide, hyaline to fuliginous. Conidiophores arising laterally on
hyaline vegetative hyphae, cylindrical with the apical part rounder, 6.75-9.65
x 2.35-3.5 um, hyaline to slightly grey. Conidiogenous cells cylindrical, with
obvious truncations at the apex, 5.9-14.4 x 2.1-3.7 um, slightly grey, attaching
directly on the terminal part of hyphae, solitary or on the conidiophores in
ageregates of 3 or more. Conidia fusoid, conspicuously attenuated from the
mid-part to the apex, slightly grey to dark fuliginous, smooth at surface,
37.55-65.15 x 4.2-7.6 um, 5-8-septate, often 7-septate, thick-walled, with
rhexolytic liberation from conidiogenous cells.

Discussion

Verticicladus hainanensis fits well within the genus Verticicladus based on
its conidiogenous cells that are verticillately fasciculate on short cylindrical
conidiophores and the multiseptate conidia with rhexolytic liberation from
conidiogenous cells.

Verticicladus hainanensis is similar to V. amazonensis, which differs by its
longer, cylindrical conidia (45-95 um; Matsushima 1993); and V. subiculifer

PiatE 1. Verticicladus hainanensis (holotype, YMF1.04024). A. Conidia; B. Conidiogenous cells
and conidia, C. Conidiophores and a conidium. Scale bars: A~C = 10 um.

IN
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278 ... Guo, Qiao, & al.

differs from V. hainensis by the different shape of its similar sized, cylindro-
fusiform conidia lacking any obvious apical attenuation (Matsushima 1996).

Acknowledgements

This work was jointly financed by National Basic Research Program of China
(973’Program: 2013CB127506) and National Natural Science Foundation Program
of PR China (31160008, 31260007, 31360130), Grants from the Young Academic and
Technical Leader Raising Foundation of Yunnan Province (2010CI020). We are very
grateful to Prof. X.G. Zhang and Dr. R.F. Castafieda-Ruiz for critically reviewing the
manuscript and providing helpful suggestions to improve this paper.

Literature cited

Ma IG, Ma J, Zhang YD, Zhang XG. 2010. A new species of Spadicoides from Yunnan, China.
Mycotaxon 113: 255-258. http://dx.doi.org/10.5248/113.255

Ma J, Wang Y, O’Neill NR, Zhang XG. 201la. A revision of the genus Lomaantha, with the
description of a new species. Mycologia 103: 407-410. http://dx.doi.org/10.3852/10-176

Ma J, Wang Y, Ma LG, Zhang YD, Castafeda-Ruiz RF, Zhang XG. 2011b. Three new species of
Neosporidesmium from Hainan, China. Mycol. Prog. 10: 157-162.
http://dx.doi.org/10.1007/s11557-010-0685-2

Ma J, Zhang YD, Ma LG, Ren SC, Castafieda-Ruiz RF, Zhang XG. 2012a. Three new species of
Solicorynespora from Hainan, China. Mycol. Progress 11: 639-645.
http://dx.doi.org/10.1007/s11557-011-0775-9

Ma LG, Ma J, Zhang YD, Zhang XG. 2012b. A new species of Corynesporella and two first records
from China. Mycotaxon 119: 83-88. http://dx.doi.org/10.5248/119.83

Matsushima T. 1993. Matsushima Mycological Memoirs, No. 7. 72 p.

Matsushima T. 1996. Matsushima Mycological Memoirs, No. 9. 30 p.

Ren SC, Ma J, Zhang XG. 2011. A new species and new records of Endophragmiella from China.
Mycotaxon 117: 123-130. http://dx.doi.org/10.5248/117.123

Ren SC, Ma J, Ma LG, Zhang YD, Zhang XG. 2012. Sativumoides and Cladosporiopsis, two new
genera of hyphomycetes from China. Mycol. Prog. 11: 443-448.
http://dx.doi.org/10.1007/s11557-011-0759-9

Wang Y, Geng Y, Ma J, Wang Q, Zhang XG. 2011. Sinomyces: a new genus of anamorphic
Pleosporaceae. Fungal Biol. 115: 188-195. http://dx.doi.org/10.1016/j.funbio.2010.12.003

Zhang K, Fu HB, Zhang XG. 2009a. Taxonomic studies of Corynespora from Hainan, China.
Mycotaxon 109: 85-93. http://dx.doi.org/10.5248/109.85

Zhang K, Fu HB, Zhang XG. 2009b. Taxonomic studies of Minimelanolocus from Yunnan, China.
Mycotaxon 109: 95-101. http://dx.doi.org/10.5248/109.95

Zhang K, Ma LG, Zhang XG. 2009c. New species and records of Shrungabeeja from southern
China. Mycologia 101: 573-578. http://dx.doi.org/10.3852/09-006

Zhang YD, Ma J, Wang Y, Ma LG, Castafieda-Ruiz RE, Zhang XG. 2011. New species and record of
Pseudoacrodictys from southern China. Mycol. Progress 10: 261-265.
http://dx.doi.org/10.1007/s11557-010-0696-z

Zhang YD, Ma J, Ma LG, Zhang XG. 2012. Two new species of Taeniolina from southern China.
Mycol. Prog. 11: 71-74. http://dx.doi.org/10.1007/s11557-010-0729-7

MY COTAXON

http://dx.doi.org/10.5248/130.279
Volume 130, pp. 279-287 January-March 2015

Geographic distribution of Sarcoporia polyspora
and S. longitubulata sp. nov.

JOSEF VLASAK™, JOSEF VLASAK JR.,
JUHA KINNUNEN’, & VIACHESLAV SPIRIN?

‘Biol. Centre of the Academy of Sciences of the Czech Republic,
Branisovskd 31, CZ-370 05 Ceské Budéjovice, Czech Rep.
*Botanical Museum, P.O. Box 7, FI-00014 University of Helsinki, Finland

* CORRESPONDENCE TO: vlasak@umbr.cas.cz

Asstract — DNA study of Sarcoporia polyspora (= Parmastomyces transmutans) revealed
only negligible sequence differences between conifer-dwelling specimens with cartilaginous
layer in the context from USA, Brazil, Europe, and Far East Asia, but a very different sequence
from three resupinate and hardwood-bound collections without such a layer and with slightly
narrower and pale brown spores from USA and Madeira Islands. This fungus, found also
among historical USA collections of S. polyspora in the BPI herbarium, is described here as
Sarcoporia longitubulata. The phylogenetic position of Sarcoporia is discussed.

Key worps — Basidiomycota, brown rot fungi, molecular taxonomy, Parmastomyces
kravtzevianus

Introduction

Sarcoporia polyspora P. Karst. is a very distinct, brown-rot polypore with
soft, resupinate to effused-reflexed basidiomes that are white to créme at first
but turn reddish-brown after bruising or drying. It can be easily recognized
by its ellipsoid and thick-walled, dextrinoid spores, which are quite unique
in polypores. The species is rare in Europe but rather common in North
America (on Tsuga spp. and Pinus spp.) and in Asia (Vlasak & Kout 2010,
Dai 2012). The fungus was described by Karsten (1894) but Karsten’s name
was abandoned for years. Based on collections from Estonia and Siberia, it
was independently described by Parmasto (1957) as Tyromyces kravtzevianus
Bondartsev & Parmasto, and a few years later Kotlaba & Pouzar (1964)
created the genus Parmastomyces for this unique polypore. In the USA,
Overholts (1941) described this species (invalidly, without a Latin diagnosis) as
“Polyporus subcartilagineus.” Later he described still another species, Polyporus

280 ... Vlasdk & al.

transmutans Oveth., which he distinguished by the more distinctly pileate habit
and its growth on Prunus (Overholts 1952). Ryvarden & Gilbertson (1984),
who found the type of P. transmutans identical with “P. subcartilagineus” as well
as with European specimens of Parmastomyces kravtzevianus, coined the name
Parmastomyces transmutans (Overh.) Ryvarden & Gilb. for all collections with
typical spores. Niemela et al. (2005), however, studied Karsten’s type material
and re-established the oldest name, Sarcoporia polyspora, for this species.

Somewhat aberrant morphology of some recent collections from hardwood
hosts and conifers led us to question the identity of Eurasian/American and
of hardwood/conifer-inhabiting Sarcoporia. We have sampled European,
American, and Asian collections from conifers and hardwoods and also
studied the type of Polyporus transmutans from the BPI herbarium along with
several other American BPI collections determined as P. transmutans to explore
whether or not all these specimens represent the same species.

Materials & methods

Morphological study

The studied specimens are deposited in the US National Fungus collection, Beltsville,
USA (BPI), in private herbarium of J. Vlasak (JV), and in mycological herbarium of
Finnish Museum of Natural History, University of Helsinki, Finland (H). They were
examined from slides prepared in cotton blue (CB) and Melzer’s reagent (IKI) with a
Leitz Diaplan microscope (x1250 magnification). Measurements were done in cotton
blue using phase contrast illumination and oil immersion (with a subjective accuracy of
0.1 um; Miettinen et al. 2006). For presenting the size range of basidiospores, 5% of the
measurements were excluded from each end of the range and are given in parentheses.
The following abbreviations are used: L’ = basidiospore length, L = mean basidiospore
length (arithmetical average of all basidiospores), W’ = basidiospore width, W = mean
basidiospore width (arithmetical average of all basidiospores), Q = L’/W’ ratio range,
and n = the number of basidiospores measured from given number of specimens. KOH-
indicates unchanged in 5% KOH and IKI- indicates unchanged in Melzer’s reagent.

Molecular phylogeny

DNA isolation and nrDNA ITS region sequencing methods follow Vlasak & Kout
(2011). The sequences were aligned with Clustal X and manually pruned. Evolutionary
analyses were conducted in MEGA6 (Tamura et al. 2013) using “all sites” and “uniform
rates” options. Other options, such as “complete deletion” and “gamma distributed with
invariant sites,” and all their combinations provided virtually identical phylogeny.

Results

Phylogenetic analysis
Sarcoporia specimens collected on hardwoods and conifers in the USA,
Madeira Island, Czech Republic, and European and Far East Russia were

Sarcoporia longitubulata sp. nov. (Macaronesia & U.S.A.) ... 281

KC595953* S. polyspora FINLAND
KC585392* P. transmutans TYPE
KC585393* S. polyspora USA conifer
KF185094* BRAZIL

Spirin 5201 RUSSIA Pinus

Spirin 4420 RUSSIA Picea

Spirin 5422 RUSSIA Picea

Spirin 6021 RUSSIA Picea

Miettinen 14814 USA Tsuga

JV 0909/1 CZECH Picea

JV 0309/93 USA Pinus

Spirin 6018 RUSSIA Pinus

JV 1009/9A hapl1 USA Quercus
JV 1009/9A hapl2 USA Quercus
100 | JV 0809/8 USA Quercus
Kinnunen 5837 MADEIRA
JQ358796* Amylocystis lapponica

Sarcoporia polyspora 100

itubulata

Sarcoporia |

0.01

FicurE 1: Phylogenetic relationships of 15 Sarcoporia specimens inferred from ITS rRNA
sequences. Amylocystis lapponica was used as outgroup. Topology from maximum likelihood
(ML) analysis. Support values along branches from ML bootstrap (1000 replicates). Branch lengths
are drawn proportional to the number of substitutions per site. GenBank numbers with asterisk
indicate sequences retrieved from GenBank; other GenBank numbers in Specimens examined.

sequenced and compared with Sarcoporia sequences in GenBank. A total
of 14 ITS and 2 nucLSU sequences were newly generated for this study and
deposited in GenBank. The ITS dataset comprising 16 sequences resulted in
an alignment with 653 characters, of which 81 were variable and 30 parsimony
informative. Extremely high sequence conservation could be demonstrated
among all conifer-dwelling specimens from different continents, with Eurasian
specimens differing from American in only one base insertion in ITS1 and
some specimens showing two haplotypes, with and without this insertion (not
shown in Fic. 1). On the other hand, most of hardwood-dwelling specimens
showed a very different sequence with about 40 mutations in ITS region.
Similarly large sequence differences were found with the nucLSU sequence
dataset (not shown), using only two sequences from each species. The aberrant
hardwood specimens were also characterized by very thick and soft resupinate
basidiocarps with long tubes and thin subiculum without cartilaginous layer.
They are described below as a new species, S. longitubulata.

282 ... Vlasdk & al.
Taxonomy

Sarcoporia longitubulata Vlasak & Spirin, sp. nov. FIGs 2- 4
MycoBank MB 809624
Sarcoporiae polysporae similis, sed tubulis longis et sporis coloratis.

Type: USA, Pennsylvania: Phoenixville, Schuylkill Canal, Ravine Trail, log of Quercus,
2 Sep 2008, J. Vlasak JV 0809/8 (Holotype, BPI 892956; isotypes, JV, H; GenBank,
KM207860, KM207863).

ErymMo oey: longitubulata (Lat.), referring to the very long tubes found in most of the

collected specimens.
BASIDIOMES annual, resupinate, pulvinaceous, up to 20 cm long, 15 cm wide,
and 2 cm thick, strikingly soft to downy, drying brittle; margin whitish to
creme, about 1 mm broad, soft and cottony, later indistinct; pore surface at
first creme with reddish tints, distinctly reddish-brown after bruising, brown to
blackish-brown on drying, the pores circular, with thin dissepiments, 3-5 per
mm; context 1-3 mm thick, soft and cottony, persistently creme colored; tube
layer very thick, up to 20 mm, extremely brittle and shattering easily when dry,
créme at first but deep brown when dry and contrasting with creme subiculum.
Spore print brown.

HYPHAL STRUCTURE monomitic; hyphae with clamp connections, KOH-,

IKI-. Context. Hyphae arranged in subparallel bundles, mostly thin-walled,
4-10 um in diam., some hyphae with thick-walls and refractive, hyaline or pale

9806
ZO

FIGURE 2: Sarcoporia basidiospores: above - S. longitubulata (holotype);
below - S. polyspora (lectotype). Scale bar = 5 um.

Sarcoporia longitubulata sp. nov. (Macaronesia & U.S.A.) ... 283

FiGurRE 3: Sarcoporia longitubulata (JV 0809/8, holotype), underside of oak log, photo in situ.
Spore deposits below left.

brown, with cyanophilous content, strongly inflated at septa, 7-18(-22) um
in diam., occasionally with double clamps. TuBEs. Hyphae subparallel, rather
loosely arranged, thin-walled, hyaline, easily collapsing, some with cyanophilous
content, (3.5-)4-5(-6) um in diam. Cystidia absent. BAsip1osporgs thick-
walled, brownish, oblong-ellipsoid, ventral side slightly concave or rarely flat,
dextrinoid, a few spores weakly or moderately cyanophilous, (4.3-)4.4-6.2
(-7.2) x (2.3-)2.5-3.3(-3.5) um (n = 150/5), L=5.29, W = 2.88, Q = 1.78-1.93.
DISTRIBUTION & ECOLOGY. On hardwood logs (predominantly Quercus

spp.) in the US North-East (Michigan, New York, Pennsylvania, Tennessee)
and Macaronesia (Madeira Is.); causing a brown rot.

ADDITIONAL SPECIMENS EXAMINED: PORTUGAL. Maperra: Madeira Island, 3 km

W from Funchal, fallen angiosperm logs, 19 Jan 2011, Kinnunen 5837 (H, GenBank

KM207865), 5845, 5848 (H). USA. PENNSYLVANIA: Gettysburg, Big Round Top,

Quercus, 5 Sep 2010, Vlasak JV 1009/9A (JV, H, GenBank KM207861, KM207862,

KM207864). Micu1GAn: Ann Arbor, hardwood, Jul 1958, Wehmeyer (BPI 223824).

NEw York: Rye, Quercus, 30 Oct 1980, Stein (BPI 222859). TENNESSEE: Great Smoky

Mt., New Found Gap, Fagus, 9 Sep 2005, Vlasak JV 0509/103 (JV, H).
ComMeEnts. Macroscopically, S. longitubulata is very distinct, so that we felt
certain from first glance that it must be a different species (Fics 3, 4). The
brown spore deposits around the basidiocarp, the very long and soft tubes,
and the relatively thin subiculum lacking a cartilaginous layer are its most
striking morphological features. Sarcoporia polyspora produces white spore
deposits and more or less pileate basidiocarps with short tubes and a thick
subiculum containing a distinct cartilaginous layer. We noted no odor of fresh

284 ... Vlasdk & al.

Figure 4: Sarcoporia longitubulata (JV 0809/8, holotype), underside of oak log, photo in situ.

basidiocarps, but on drying, S. longitubulata smelled so repugnantly that it
was impossible to keep in the same room for more than one week, although
it was not contaminated with yeasts. The primary microscopic difference of
S. longitubulata is the pale brown coloration of the basidiospores, visible in
cotton blue even with low resolution; only a few spores are more or less stained
(slightly or moderately cyanophilous). In contrast, S. polyspora basidiospores
are exceptionally strongly cyanophilous (uniformly bright blue), so that their
natural color is totally undetectable in cotton blue. Moreover, the S. longitubulata
spores are a bit narrower than in S. polyspora (TaBLE 1) and less variable in
shape and size, being uniformly oblong-ellipsoid and slightly concave on their
ventral side (Fic. 2). The basidiospores of this type occur in S. polyspora, too,
but only as a rare variation.

Discussion

We have shown that S. polyspora is widely distributed from America to
Europe and Far East Asia. The slight morphological differences we observed—
e.g., longer spores and more pileate habit in USA collections vs. shorter spores
in Far East Asian collections—have no support in DNA sequence differences
and so should be regarded as regional morphological variation. A tropical
distribution remains uncertain, as the sequenced specimens from Brazil were
collected from Pinus logs probably imported from the USA (Baldoni et al.
2015).

All studied collections were from conifers except for BPI 844703, the type of
Polyporus transmutans (Overholts 1952), collected on Prunus serotina Ehrh. in
northwestern USA. This specimen displays strikingly large and tough pilei with

Sarcoporia longitubulata sp. nov. (Macaronesia & U.S.A.) ... 285

TABLE 1. Spore measurements of two Sarcoporia species. (Limit and mean values for
each species shown in bold.)

SPECIES / SPECIMEN Ly L W’ W Q n

S. polyspora (4.2)4.7-7.2(7.3) 5.53 (2.5)2.6-3.8(4.0) 3.10 1.79 270
Miettinen 14814 (5.0)5.1-6.3(6.7) 5.64 (2.7)2.8-3.6(4.2) 3.14 1.81 30
Spirin 4420 (4.9)5.0-6.3(6.7) 5.51 (2.7)2.8-3.2(3.3) 2.97 1.86 30
Spirin 5201 (4.2)4.7-5.7(5.8) 5.05 (2.6)2.7-3.3 3.02 1.68 30
Spirin 5422 (4.7)4.8-5.7(7.1) 5.22 (2.5)2.6-3.2(3.3) 2.93 1.79 30
Spirin 6021 (4.8)5.2-6.8(7.3) 5.70 (2.6)2.7-3.6(3.7) 3.07 1.86 30
Spirin 6018 (4.8)4.9-6.1(6.2) 5.30 (2.7)2.8-3.4(3.7) 3.11 1.71 30
Vlasak 0908/15 (4.8)5.0-6.2(6.4) 5.56 (2.7)2.8-3.7(3.9) 3.17 1.76 30
Vlasak 0309/93 (5.2)5.3-7.2(7.3) 6.27 (2.8)2.9-3.8(4.0) 3.19 1.97 30

S. longitubulata (4.3)4.4-6.2(7.2) 5.29 (2.3)2.5-3.3(3.5) 2.88 1.84 150
Vlasak 0809/8 (4.3)4.4-5.8(6.0) 5.03 (2.6)2.7-3.1(3.3) 2.82 1.79 30
Vlasak 1009/9a (4.6)4.8-6.2(6.3) 5.42 (2.3)2.5-3.2(3.3) 2.82 1.93 30
Vlasak 0509/103 (4.4)4.8-5.8(7.2) 5.30 (2.7)2.8-3.3(3.5) 2.97 1.78 30
BPI 222859 (4.8)4.9-6.2(6.7) 5.39 (2.6)2.7-3.2(3.3) 2.90 1.86 30
BPI 223824 (4.8)4.9-6.0 5.32 (2.5)2.6-3.3(3.4) 2.91 1.83 30

short tubes and thick context in which no distinct gelatinous layer is present
but scattered gelatinous streaks are present. This suggests just a very large
specimen of S. polyspora and our examination revealed no distinct microscopic
differences from other S. polyspora specimens. Moreover, the ITS sequence
of the culture derived from this specimen available in GenBank (KC585392;
Ortiz-Santana et al. 2013) is identical with other S. polyspora sequences.

When we sought for a possible existing name for S. longitubulata, Tyromyces
mollissimus Maire, described from North Africa (Maire 1945) and presently
regarded as a synonym of S. polyspora, seemed a reasonable candidate.
Unfortunately, the type material is now unavailable (Pouzar 1984, Kotiranta
1998). Nevertheless, the protologue of T’ mollissimus is very clear and leaves
no doubts about its identity: it was described as growing on Pinus halepensis
Mill. and having pileate or effused-reflexed, very soft basidiocarps with thick
contextual layer (see fig. 7b, Maire 1945: 37), and a white spore print. All these
features fit well with the current concept of S. polyspora and certainly rule out
our new species. The recently described Sarcoporia neotropica from Costa Rica
(Ryvarden 2013) has imbricate soft basidiocarps and hyaline basidiospores,
characters that preclude its identity with S. longitubulata.

Because of similar spores and some of tissue features, Ryvarden &
Melo (2014) speculated about Sarcoporia’s position in the Coniophoraceae.
Molecular phylogeny refutes this hypothesis, however. The sequences of typical

286 ... Vlasak & al.

coniophoraceous species such as Coniophora puteana (Schumach.) P. Karst.
and Serpula lacrymans (Wulfen) J. Schrot. differ greatly from Sarcoporia,
which has already been recognized as a member of “antrodia clade” within
the Polyporaceae, most closely related to Amylocystis and Auriporia (Ortiz-
Santana et al. 2013). Amylocystis lapponica (Romell) Bondartsev & Singer,
is in fact quite similar, being soft and turning reddish brown after bruising
and drying, causing a brown rot, and with some microstructures staining in
Melzer’s reagent. Bondarcevomyces taxi (Bondartsev) Parmasto, which was
once referred to Parmastomyces (= Sarcoporia in current sense) by Dai &
Niemela (1995), belongs to the Boletales (Binder & Hibbett 2006). It does share
some characters with S. longitubulata (brownish spores, strong pungent odor of
drying basidiocarps) although they indicate only superficial similarity.
ADDITIONAL MATERIAL EXAMINED: Sarcoporia polyspora. CZECH REPUBLIC.
Hluboka, Karluv Hradek, Picea abies, 2 Sep 2009 Vlasak JV 0909/1 (JV, GenBank
KM207858, KM207859), 11 Aug 2009, Vlasak JV 0908/15 (JV, PRM 915664, H).
FINLAND. Uusimaa: Inkoo, Fagervik, Sept 1893, Hisinger (H) (lectotype of S. polyspora;
Lowe 1956). RUSSIA. KHABAROVSK REG.: Khabarovsk Dist., Ulika, Pinus koraiensis, 13
Aug 2012, Spirin 5201 (H, GenBank KM207856, KM207857), Malyi Kukachan, Picea
ajanensis, 19 Aug 2012, Spirin 5422 (H, GenBank KM207852); Solnechnyi Dist., Evoron,
Picea obovata, 27 Aug 2011, Spirin 4420 (H, GenBank KM207853). NIZHNY NOVGOROD
REG.: Lukoyanov Dist., Panzelka, Pinus sylvestris, 8 Aug 2013, Spirin 6018 (H, GenBank
KM207851), Picea abies, 8 Aug 2013, Spirin 6021 (H, GenBank KM207854, KM207855).
USA. MassacuusetTts: Holden, Quinapoxet, Tsuga canadensis, 26 Sep 2011, Miettinen
14814.1 (H, GenBank KM207850). PENNSYLVANIA: Wilkes-Barre, Ricketts Glen St.
Park, Pinus, 11 Sep 2003, Vlasak JV 0309/93 (JV, H, GenBank KM207849); Haycock,
Nockamixon St. Park, Pinus strobus, 7 Aug 2004, Vlasak Jr. JV 0408/4) (JV); McKean
Co., Prunus serotina, 10 Sep 1940, Campbell & Davidson 22971 (BPI 844703, holotype
of Polyporus transmutans, GenBank KC585392). NEw York: Adirondack Park, Keene
Valley, Johns Brook Trail, conifer, 23 Sep 2005, Vlasak Jr. JV 0509/193J (JV).
Bondarcevomyces taxi. RUSSIA. KHABAROvSK REG.: Khabarovsk Dist., Bolshoi
Khekhtsir Nat Res., Picea ajanensis, 5 Sep 2013, Spirin 6674 (H, JV); Komsomol’sk Dist.,
Boktor, Larix gmelinii, 18 Aug 2013, Spirin 6141 (H, JV).

Acknowledgments

The authors thank the BPI herbarium staff for the kind loan of Sarcoporia specimens,
Dr. Otto Miettinen for sequencing the Kinnunen 5837 specimen, and Prof. Teuvo Ahti
for revising the Latin diagnosis. Valuable discussions with Jiri Kout, G. Coelho, and
Y.-C. Dai and their contributions as presubmission reviewers are acknowledged. The
research of J. Vlasak was funded by institutional support RVO: 60077344.

Literature cited

Baldoni DB, Ortiz-Santana B, Coelho G, Antoniolli ZI, Jacques RJS. 2015. Sarcoporia polyspora
(Basidiomycota, Polyporales): a rare wood-decay fungus newly recorded from South America.
Nova Hedwigia 100: 177-187.
http://dx.doi.org.ezproxy.landcareresearch.co.nz/10.1127/nova_hedwigia/2014/0218

Sarcoporia longitubulata sp. nov. (Macaronesia & U.S.A.) ... 287

Binder M, Hibbett DS. 2006. Molecular systematics and biological diversification of Boletales.
Mycologia 98(6): 971-981. http://dx.doi.org/10.3852/mycologia.98.6.971

Dai YC. 2012. Polypore diversity in China with an annotated checklist of Chinese polypores.
Mycoscience 53: 49-80. http://dx.doi.org/10.1007/s10267-011-0134-3

Dai YC, Niemela T. 1995. Changbai wood-rotting fungi 4. Some species described by A.S.
Bondartsev and L.V. Lyubarsky from the Russian Far East. Annales Botanici Fennici 32(4):
AL1-226.

Karsten PA. 1894. Fragmenta mycologica XLII. Hedwigia 33: 15-16.

Kotiranta H. 1998. Parmastomyces mollissimus in North Europe. Folia Cryptog. Estonica 33: 41-47.

Kotlaba F, Pouzar Z. 1964. Staining spores of Homobasidiomycetes in cotton blue and its importance
for taxonomy. Feddes Repert. 69: 131-142.

Maire R. 1945. Etudes mycologiques. Fascicle 5. Bull. Soc. Hist. Nat. Afr. N. 36: 24-42.

Miettinen O, Niemela T, Spirin W. 2006. Northern Antrodiella species: the identity of A. semisupina,
and type studies of related taxa. Mycotaxon 96: 211-239.

Niemela T, Kinnunen J, Larsson KH, Schigel D, Larsson E. 2005. Genus revisions and the new
combinations of some North European polypores. Karstenia 45: 75-80.

Ortiz-Santana B, Lindner DL, Miettinen O, Justo A, Hibbet DS. 2013. A phylogenetic overview of
the antrodia clade (Basidiomycota, Polyporales). Mycologia 105: 1391-1411.
http://dx.doi.org/10.3852/13-051

Overholts LO. 1941. New species of Polyporaceae. Mycologia 33: 90-102.
http://dx.doi.org/10.2307/3754741

Overholts LO. 1952. New species of polypores. Mycologia 44: 224-227.

Parmasto E. 1957. Eesti seente eksikaat. Mycotheca Estonica 1: no. 1-25. Tartu. 22 p.

Pouzar Z. 1984. Notes on four European polypores. Ceska Mykol. 38: 203-204.

Ryvarden L. 2013. Studies in neotropical polypores 32. Some new species from Costa Rica. Synopsis
Fungorum 30: 33-43.

Ryvarden L, Gilbertson RL. 1984. Type studies in the Polyporaceae 15, species described by O.L.
Overholts, either alone or with J.L. Lowe. Mycotaxon 19: 137-144.

Ryvarden L, Melo I. 2014. Poroid fungi of Europe. Synopsis Fungorum 31: 1-455

Tamura K, Stecher G, Peterson D, Filipski A, Kumar S. 2013. MEGA 6: Molecular Evolutionary
Genetics Analysis version 6.0. Mol. Biol. Evol. 30: 2725-2729.
http://dx.doi.org/10.1093/molbev/mst197

Vlasak J, Kout J. 2010. Sarcoporia polyspora and Jahnoporus hirtus: two rare polypores collected in
South Bohemia, Czech Republic. Czech Mycol. 61(2): 187-195.

Vlasak J, Kout J. 2011. Tropical Trametes lactinea is widely distributed in the eastern USA.
Mycotaxon 115: 271-279. http://dx.doi.org/10.5248/115.271

MY COTAXON

http://dx.doi.org/10.5248/130.289
Volume 130, pp. 289-293 January-March 2015

Coccomyces neolitseae sp. nov. from Zhangjiajie, China

QING LI’, YU-XIA CHEN’, CHUN-TAO ZHENG’,
DAN-DAN Lu! & YING-REN LIN”

' School of Life Science &? School of Forestry & Landscape Architecture,
Anhui Agricultural University, West Changjiang Road 130, Hefei, Anhui 230036, China

*CORRESPONDENCE TO: yingrenlin@yahoo.com

Asstract — A fungus found on leaves of Neolitsea levinei from the Zhangjiajie National
Forest Park in Hunan Province, China, is described as Coccomyces neolitseae. A description,
illustration, and comments are given for this taxon. The type specimen is deposited in the
Reference Collection of Forest Fungi of Anhui Agricultural University, China (AAUF).

Key worps — Rhytismataceae, Rhytismatales, morphological characteristics, taxonomy,
Lauraceae

Introduction

Coccomyces De Not. is now a large genus in the Rhytismataceae (Kirk et al.
2008). The ascomal external shape, size, and type of opening, the ascal shape
and size, and the shape of paraphyses at the apex are regarded as the most
important taxonomic characters of Coccomyces (Johnston 1986, Lin 1998).
Index Fungorum (2015) lists 196 epithets for the genus, of which Species
Fungorum (2015) accepts 106 species in Coccomyces. They usually occur
on leaves, herbaceous stems, bark, and wood of vascular plants, especially
Ericaceae, Fagaceae, and Lauraceae (Sherwood 1980). Some species, such as
C. guizhouensis Y.R. Lin & B.E. Hu, C. ledi Rehm, C. strobi J. Reid & Cain, and
C. vilis Syd. et al., may cause economically significant plant diseases (Lin et al.
1994, Rehm 1913, Reid & Cain 1961, Cannon & Minter 1984).

Recently, another new Coccomyces species was found on fallen leaves of
Neolitsea levinei in the Zhangijiajie National Forest Park, Hunan Province,
which we describe below.

Materials & methods
The study was based on collections made in 2010 from Hunan, China. Specimens
were rehydrated in water for 15 min and ascomata and conidiomata were sliced into 10-

290 ... Li & al.

15 um thick sections using a freezing microtome. The structure of the fruiting bodies
was observed in 0.1% (w/v) cotton blue in phenol glycerin. The color of the internal
structures of the fruiting bodies was observed in water. Measurements and drawings of
asci, ascospores, and paraphyses were made from squash mounts in 5% KOH solution.

Taxonomy

Coccomyces neolitseae Qing Li & Y.R. Lin, sp. nov. Fics 1-6
MycoBAnk MB 809422
Differs from Coccomyces delta by the lack of lip cells, the existence of an excipulum, a
thicker basal stroma, paraphyses that are not enlarged at the apex, shorter and thinner
ascospores lacking an obvious gelatinous sheath, and the ascomata associating with
conidiomata.

Type: China, Hunan, Zhangjiajie National Forest Park, Jinbianxi, on fallen leaves
of Neolitsea levinei Merr. (Lauraceae), 17 August 2010, X.M. Gao & Y.R. Lin 2546
(Holotype, AAUF 68654).

Erymo oey: The specific epithet refers to the host genus.

Cotoniegs on both sides of leaves, forming irregular, yellowish-white spots
6-20 mm diam.

ZONE LINES frequent, dark-brown or black, thin, entirely or partly
surrounding the bleached areas.

ConipiomaTa on both sides of leaves, predominantly on the lower side,
scattered to clustered, sometimes with several coalescing. In surface view,
conidiomata 110-215(-250) um diam., subcircular, brown to black-brown,
with a marked perimeter line or clearly defined edge, somewhat raising the
substratum surface, discharging spores through an apical ostiole. In vertical
transverse section, conidiomata intraepidermal, more or less lens-shaped.
UPPER WALL 3.5-6.5 um thick, dark brown, composed of tiny, thin-walled
angular cells 1-1.5 um diam. BAasAL WALL 2.5-6 um thick, comprised of
(1-)2-3 layers of thick-walled angular cells 2-3 um diam. SUBCONIDIOGENOUS
LAYER 2.5-4 um thick, consisting of nearly colorless, thin-walled angular cells.
CONIDIOGENOUS CELLS and ConrIpIA not present in material available.

Ascomarta in similar positions to conidiomata on the substratum, scattered,
occasionally two coalescing in the bleached spots. In surface view, ascomata
800-1200(-1600) um diam., triangular, occasionally quadrilateral, black,
with a distinct outline, moderately raising the substratum surface,
with an obvious preformed dehiscence mechanism, opening by 3-4 radial
splits that extend nearly to the ascomatal edge to expose the light yellow-
brown hymenium. Lips absent. In median vertical transverse section, ascomata
intraepidermal. COVERING STROMA 45-55 um thick in the middle of the
laterals, becoming thinner towards the top and the edge, connecting to the basal
stroma, composed of black-brown textura angularis with thick-walled cells 6-8

Coccomyces neolitseae sp. nov. (China) ... 291

Fics 1-6. Coccomyces neolitseae (holotype, AAUF 68654) on Neolitsea levinei. 1. Habit on a leaf.
2. Detail of conidiomata, ascomata, and a zone line. 3. Ascoma in median vertical transverse
section. 4. Conidioma in vertical transverse section. 5. Portion of ascoma in median transverse
vertical section. 6. Paraphyses, asci, and ascospores.

292-0 eal.

um diam. BASAL STROMA 10-18 um thick, consisting of textura angularis with
2-3 layers of black-brown, thick-walled cells. INTERNAL MATRIX OF STROMA
existing only between covering stroma and basal stroma, composed of colorless,
thin-walled angular cells 7-12 um diam., Excipu_um well-developed, 25-35
uum wide, arising from the edge of the subhymenium, comprised of rows of
hyaline, thin-walled, septate hyphae. SUBHYMENIUM 12-20 um thick, composed
of colorless textura porrecta-intricata. PARAPHYSES hyaline, aseptate, 120-150
x 0.8-1.4 um, filiform, sinuate, not swollen, occasionally branched at the apex,
not immersed in a gel. Asci ripening sequentially, 105-120 x 8-11 um, clavate,
long-stalked, thin-walled, apex rounded to obtuse, without circumapical
thickening, J-, 8-spored. AscosporEs arranged in a fascicle, 64-75 x 1.5-1.7
um, filiform, hyaline, aseptate, gradually tapered towards the acute base, not
obviously sheathed.

HOST SPECIES, HABITAT, & DISTRIBUTION: Producing conidiomata and
ascomata on fallen leaves of Neolitsea levinei; known only from the type locality,
Hunan Province, China.

COMMENTS — Coccomyces delta (Kunze) Sacc. is closely related to C. neolitseae in
ascomal appearance and in the way it is embedded, as well as the characteristics
of the zone lines. However, C. delta differs in the lack of an association of its
ascomata with conidiomata, the absence of an excipulum, the presence of a
fringe of periphysis-like lip cells, a much thinner (5 um thick) basal stroma,
slightly enlarged paraphysial apices, short-stalked, longer (100-150 tm) asci,
and wider and longer (80-100 x 2 um) ascospores with a narrow mucous
sheath (Sherwood 1980). Coccomyces delta has been observed on a wide range
of woody plants, mostly in the Europe and Atlantic islands, but with some
records from China (Sherwood 1980; Lin et al. 2012). In our Chinese materials,
the paraphyses are septate and covered by a thin mucous coating, and the asci
and ascospores are shorter than described by Sherwood (1980).

Coccomyces limitatus (Berk. & M.A. Curtis) Sacc. is easily distinguished
from the new species by trilateral to pentagonal ascomata, a covering stroma
that does not extend as far as the basal stroma, a well-developed internal
matrix comprising loose hyphae and crystals, a wider (50 um thick) excipulum
arising from inner layers of upper wall, wider (2-3 um wide) paraphyses with
swollen upper apices, and dissimilar asci and ascospores. Its asci are 4-5.6 um,
sometimes with an apical thickening, and the ascospores are only 0.8-1 um
wide and do not taper to the base (Johnston 1986).

Coccomyces neolitseae resembles Coccomyces huangshanensis Y.R. Lin & Z.Z. Li
in the nature of the internal matrix of stroma and the color of the exposed
hymenium, but C. huangshanensis has a much thinner covering stroma (8-17

Coccomyces neolitseae sp. nov. (China) ... 293

um) covered by heavily carbonized tissue at the opening that thins gradually
towards the ascomal edge, short and hyaline periphysoids, and ascospores
sometimes with a gelatinous cap at the apex (Lin et al. 2000).

Acknowledgements

The authors are grateful to Dr J.E. Taylor (Royal Botanic Garden Edinburgh, UK)
and Dr M. Ye (Hefei University of Technology, China) for serving as pre-submission
reviewers and to the Dr L.H. Wang for the identification of host plant. This study
was supported by the National Natural Science Foundation of China (No. 31270065,
31170019).

Literature cited

Cannon PF, Minter DW. 1984. Coccomyces vilis. CMI Descr. Path. Fungi & Bact. no. 792.

Index Fungorum. 2015. www.indexfungorum.org [viewed online on 23 March 2015].

Johnston PR. 1986. Rhytismataceae in New Zealand 1. Some foliicolous species of Coccomyces de
Notaris and Propolis (Fries) Corda. New Zealand J. Bot. 24: 89-124. http://dx.doi.org/10.1080
/0028825X.1986.10409723

Kirk PM, Cannon PF, Minter DW, Stalpers JA. 2008. Ainsworth & Bisby’s dictionary of the fungi,
10" ed. CAB International. Wallingford. 771 p.

Lin YR. 1998. Studies on Coccomyces de Notaris and Neococcomyces gen. nov. in China. [Ph.D.
thesis; in Chinese]. Northeast Forestry University. Harbin. 98 p.

Lin YR, Liu HY, Tang YP, Hu BE. 1994. Two new species of the genus Coccomyces from China [in
Chinese]. Acta Mycol. Sin. 13: 8-12.

Lin YR, Li ZZ, Xie YS, Liang SW. 2000. Studies on the genus Coccomyces from China III [in
Chinese]. Mycosystema 19: 449-453.

Lin YR, Liu HY, Hou CL, Wang SJ, Ye M, Huang CL, Xiang Y, Yu SM. 2012. Flora Fungorum
Sinicorum, vol. 40, Rhytismatales [in Chinese]. Science Press. Beijing. 261 p.

Rehm H. 1913. Ascomycetes novi VI. Ann. Mycol. 11: 150-155.

Reid J, Cain RE. 1961. The genus Therrya. Can J. Bot. 39: 1119-1129.
http://dx.doi.org/10.1139/b61-098

Sherwood MA. 1980. Taxonomic studies in the Phacidiales: The genus Coccomyces (Rhytismataceae).
Occ. Pap. Farlow Herb. Crypt. Bot. 15: 1-120.

Species Fungorum. 2015. www.speciesfungorum.org [viewed online on 23 March 2015].

MY COTAXON

http://dx.doi.org/10.5248/130.295
Volume 130, pp. 295-297 January-March 2015

Datronia ustulatiligna sp. nov. (Agaricomycetes) from India

HARPREET KAUR, GURPREET KAUR & G.S. DHINGRA

Department of Botany, Punjabi University, Patiala 147 002, India

* CORRESPONDENCE TO: harpreetkaur153@gmail.com

ABSTRACT — A new poroid species, Datronia ustulatiligna, is described on partly burned
gymnospermous wood from the state of Himachal Pradesh in India.

Key worps — Basidiomycota, Rampur, Habban

While conducting fungal forays in the Narkanda, Rampur, Rajgarh, and
Habban areas of the Shimla and Sirmaur districts in Himachal Pradesh (India),
Harpreet and Dhingra made some collections of an unknown poroid fungus
associated with partly burned trees of Pinus roxburghii and Cedrus deodara.
After comparing macroscopic and microscopic features with published
descriptions (Bakshi 1971, Ryvarden & Gilbertson 1993, Nunez & Ryvarden
2001, Bernicchia 2004, Sharma 2012), we identified the polypore as a new
species of Datronia close to D. scutellata. It is distinguished by a combination of
its association with partly burned gymnospermous wood, larger pores (2-4 per
mm), and long narrowly clavate basidia.

Datronia ustulatiligna Har. Kaur, G. Kaur & Dhingra, sp. nov. PLATE 1
MycoBank MB 810285
Differs from Datronia scutellata in having thicker basidiocarps, larger pores, and long,

narrowly clavate basidia.

Type: India, Himachal Pradesh, Shimla, 8 km from Narkanda towards Rampur, on a
partly burned tree of Pinus roxburghii Sarg., 2 August 2010, Harpreet Kaur 5609 (PUN,
holotype).

Erymo.ocy: The epithet refers to the substrate, partly burned wood.

Basidiocarps annual to biennial, pileate, effused-reflexed to sessile, solitary
to imbricate; up to 4 cm wide, 2.2 cm in radius, 2.2 cm thick near base,
applanate to ungulate to dimidiate; abhymenial surface scrupose to somewhat
hirsute, concentrically sulcate and zonate, often covered with mosses, brown

296 ... Kaur, Kaur, & Dhingra

8 12

PLATE 1. Datronia ustulatiligna (holotype). 1. Hymenophore (fresh). 2. Upper part of pileus (fresh).
3. Upper part of pileus covered with mosses (fresh). 4. Hymenophore (dry). 5. Upper part of pileus
(dry). 6. Pores (with mm scale). 7. Basidiospores. 8. Basidia. 9. Groups of crystals. 10. Generative
hyphae. 11. Cystidioles. 12. Skeletal hyphae.

Datronia ustulatiligna sp. nov. (India) ... 297

to dark brown when fresh, do not change much on drying; hymenial surface
poroid, light brown to brown when fresh, brown to dark brown on drying;
pores angular to daedaleoid, 2-4 per mm; dissepiments thick to thin, entire
to lacerate; context <2 mm thick, somewhat duplex near base, brown; pore
tubes <2.0 mm long, brown; scattered groups of crystals observed in trama;
margins acute to obtuse, regular to lobed, sterile <1 mm, concolorous. Hyphal
system dimitic. Generative hyphae <3.3 um wide, branched, septate, clamped,
thin-walled in subhymenium and thick-walled in context, subhyaline. Skeletal
hyphae <3.3 um wide, rarely branched, aseptate, thick-walled, yellowish brown.
Cystidia absent, but cystidioles present in hymenial layer, 16-37 x 4-5.3 um,
fusoid, thin-walled. Basidia 40-49 x 4-6 um, narrowly clavate, 4-sterigmate,
subhyaline, clamped at the base; sterigmata up to 3.3 um long. Basidiospores
7.3-9.8 x 2.5-4 um, ellipsoid to subcylindrical to somewhat allantoid, smooth,
subhyaline, with oily contents, thin-walled inamyloid, acyanophilous.
ADDITIONAL SPECIMENS EXAMINED: INDIA, HIMACHAL PRADESH, Shimla, about 4 km
from Narkanda towards Rampur, 2 August 2010, on a partly burned gymnospermous
stump, Dhingra 5611 (PUN); on a partly burned tree of P. roxburghii, Harpreet 5608
(PUN), Harpreet 5613 (PUN); Sirmaur, 4 km from Habban towards Rajgarh, 30 August
2011, ona partly burned tree of Cedrus deodara (D. Don) G. Don, Harpreet 5612 (PUN).
REMARKS- Datronia scutellata (Schwein.) Gilb. & Ryvarden differs from our
new species by smaller pores (4-5 per mm), shorter and wider basidia (20-30
x 7-10 um), and association with angiospermous wood.

Acknowledgements

The authors thank Head (Department of Botany, Punjabi University, Patiala) for
providing research facilities; Dr. Nils Hallenberg, Professor Emeritus (University of
Gothenburg), for peer review and expert comments; and Prof. B.M. Sharma (Department
of Plant Pathology, COA, CSKHPAU, Palampur, H.P., India) for peer review.

Literature cited

Bakshi BL. 1971. Indian Polyporaceae (on trees and timber). Indian Council of Agricultural
Research. New Delhi. 246 p.

Bernicchia A. 2004. Polyporaceae s.1. Fungi Europaei. Edizioni Candusso, Alassio, Italia. 808 p.

Nufez M, Ryvarden L. 2001. East Asian polypores 2. Polyporaceae s. lato. Synopsis Fungorum 14:
170-522.

Ryvarden L, Gilbertson RL. 1993. European polypores, vol. 1. Abortiporus - Lindtneria. Fungiflora,
Oslo. pp. 1-387

Sharma JR. 2012. Aphyllophorales of Himalaya (Auriscalpiaceae-Tremellodendropsis). Botanical
Survey of India. 590 p.

MYCOTAXON

http://dx.doi.org/10.5248/130.299
Volume 130, pp. 299-304 January-March 2015

Three new species of Xylaria from China

Gu HuANG’”, RUISHA WANG', LIN Guo’, & NA Liu?

'State Key Laboratory of Mycology, Institute of Microbiology, Chinese Academy of Sciences,
Beijing 100101, China

College of Biological Engineering, Henan University of Technology, Zhengzhou 450001, China

College of Life Sciences, Hebei Normal University, Shijiazhuang 050016, China

* CORRESPONDENCE TO: guol@im.ac.cn

ABSTRACT —Three new species, Xylaria semiglobosa, X. sphaerica, and X. jiangsuensis, are
described. They were discovered in Yunnan, Hainan, and Jiangsu Provinces in China.

KEY worDs —Ascomycota, pyrenomycetous fungi, Xylariaceae, taxonomy

Teng (1963) described 51 species and one variety of Xylaria from China and
Tai (1979) recorded 52 species, one variety, and one form. Ju & Roger (1999)
reported 40 species and 1 variety from Taiwan. Since 1979 many new Chinese
species and records of Xylaria have been published (Li & Li 1994, Xu 1999, Ju
& Hsieh 2007, Ma et al. 2011a,b, 2012, 2013, Zhu & Guo 2011, Huang et al.
2014a,b). Three additional new species are now described as follows:

Xylaria semiglobosa G. Huang & L. Guo, sp. nov. Figs 1-3
FUNGAL NAME FN570116
Differs from Xylaria glebulosa in its larger stromata and smaller ascospores.
Type: China, Yunnan, Mengla, alt. 760 m, on wood, 17.X.2013, G. Huang, L. Guo & W.
Li 230 (HMAS 270193, holotype).

Erymo oey: The epithet refers to the stromatal shape.

Stromata gregarious or scattered, semiglobose, on narrow central connective,
3-9 mm high, 4-14 mm diam; surface black, finely reticulately cracked, with
inconspicuous perithecial mounds; interior white, becoming hollow at maturity.
Perithecia subglobose or ellipsoid, 600-900 um diam; ostioles papillate. Asci
with eight ascospores arranged in a uniseriate or partially biseriate manner,
cylindrical, overall 212-237 um long, 9-16 um broad, the spore-bearing parts

300 ... Huang & al.

122-132 um long, with an apical ring staining blue in Melzer’s iodine reagent,
urn-shaped, 9-10 um high, 5-8 um broad. Ascospores dark brown, unicellular,
ellipsoid-inequilateral, sometimes pinched at the one end, smooth, (20-)
22-25(-27) x 6-7(-9) um, with an oblique germ slit much shorter than the
spore length.

ADDITIONAL SPECIMEN EXAMINED: CHINA, Hatna. Diaoluoshan Natural Reserve, on

wood, 10.X1.2012, S.H. He 201211101 (HMAS 270192).
Comments: Xylaria semiglobosa is similar to X. glebulosa (Ces.) Y.M. Ju &
J.D. Rogers, which differs in having smaller stromata [(1.5-3) mm high, 1-3
mm diam] and larger ascospores (27-31 x 8-10 um), frequently with abruptly
pinched ends (Ju & Rogers 1999). The new species also resembles X. fraseri
M.A. Whalley et al., which differs in having whitish scales on the stromatal
surface, asci with a shorter apical ring (4.5-5 um high, 3.5-4 um broad), and
ascospores with a straight germ slit (Whalley et al. 2000). Xylaria semiglobosa
is close to X. atroglobosa Hai X. Ma et al., which differs in its asci with shorter
apical rings (4.5-5 um high, 3.5-5 um broad) and ascospores with a round
hyaline non-cellular appendage up to 6-7 x 3-4 um long (Ma et al. 2012).

Xylaria sphaerica G. Huang & L. Guo, sp. nov. Fics 4-6
FUNGAL NAME FN570117

Differs from Xylaria sicula f. major in its stromata on wood with a small acute sterile
apex and long branched protrusion near the base of the head and its slightly larger
ascospores.

Type: China, Hainan, Bawangling Natural Reserve, alt. 740 m, on wood, 12.XII.2009,
Y.F. Zhu & L. Guo 133 (HMAS 270191, holotype).

Erymo.oey: The epithet refers to the stromatal shape.

Stromata with small acute sterile apex and long branches protruding
near the base of the head, 1.3-1.5 cm total length, fertile parts spherical or
subsphaeroidal, 1-1.5 mm high, 1-2 mm diam; on tomentose hair-like stipes;
surface black, smooth, with perithecial mounds; interior white. Perithecia
subglobose or ellipsoid, 540-600 um diam; ostioles papillate. Asci with eight
ascospores arranged in uniseriate manner, cylindrical, 118-128 um total
length, 7-12 um broad, the spore-bearing parts 75-83 um long, with an apical
ring staining blue in Melzer’s iodine reagent, hat-shaped, 4-5 um high, 3-4
um broad. Ascospores brown, unicellular, ellipsoid—inequilateral, occasionally
pinched at the one end, smooth, (10.5-)12-13(-15) x 5-7 um, with a straight
germ slit the length of the spore.

Fics 1-6. Xylaria semiglobosa (HMAS 270193, holotype). 1. Stromata on wood; 2. Section of
stroma; 3. Ascus and ascospores. Xylaria sphaerica (HMAS 270191, holotype). 4. Stromata on
wood; 5. Section of stroma; 6. Asci and ascospores.

Xylaria spp. nov. (China) ... 301

302 ... Huang & al.

ComMENnts: Xylaria sphaerica is similar to X. sicula f. major Ciccar., which
differs in having stromata on fallen leaves and with a long apex, slightly smaller
ascospores (9-12 x 3-6 um), and the absence of protruding branches near the
base of the stromatal head (Ciccarone 1946).

Xylaria jiangsuensis Rui S. Wang & L. Guo, sp. nov. Fics 7-10
FUNGAL NAME FN570118
Differs from Xylaria filiformis in its stromata with branched stipes and its larger and
fusiform ascospores.
Type: China, Jiangsu: Baohuashan, on dead leaves, 10.V1I.1932, S.C. Teng 977 (HMAS
7263, holotype).

Erymo.oey: The epithet refers to Jiangsu Province, where this fungus was first collected.

Stromata filiform, with a sterile apex, 3-7.5 cm total length, 0.5-1 mm diam,
fertile parts 3-6 mm long; on hair-like, branched stipes; surface black, smooth,
with conspicuous perithecial mounds; interior white. Perithecia globose or
subglobose, 450-570 um diam; ostioles papillate. Asci with eight ascospores
arranged in uniseriate manner, cylindrical, the spore-bearing parts 104-110
uum long, 5-7 um broad, with an apical ring staining blue in Melzer’s iodine
reagent, inverted hat-shaped, 2-2.5 um high, 1-1.5 um broad. Ascospores
brownish or brown, unicellular, fusiform-inequilateral, sometimes pinched at
the one end, smooth, 16.5-20(-21.5) x 4-5(-6) um, with straight germ slit the
length of the spore.

ComMENTSs: Xylaria jiangsuensis is similar to X. filiformis (Alb. & Schwein.)
Fr., which differs in having stromata on unbranched stipes and its ellipsoid-
inequilateral and smaller ascospores (11-14 x 5-6 um; Rogers & Ju 1998).

Acknowledgements

The authors would like to express their deep thanks to Prof. Anthony J.S. Whalley
(Liverpool, UK) and Dr. L. Vasilyeva (Vladivostok, Russia) for serving as pre-submission
reviewers and to Dr. Shaun Pennycook (Auckland, New Zealand) for nomenclatural
review. This study was supported by the foundation of Project of the Knowledge
Innovation Program of the Chinese Academy of Sciences (No. KSCX2-EW-Z-9).

Literature cited

Ciccarone A. 1946. Alcune osservazioni su una forma di Xylaria sicula Pass. e Beltr. Nuovo Giorn.
Bot. Ital., n.s. 53: 356-358.

Huang G, Guo L, Liu N. 2014a. Xylaria byttneriae sp. nov. from Yunnan Province in China.
Mycosystema 33: 567-570.

Huang G, Guo L, Liu N. 2014b.Two new species of Xylaria and X. diminuta new to China.
Mycotaxon 129(1): 149-152. http://dx.doi.org/10.5248/129.149

Ju YM, Rogers JD. 1999. The Xylariaceae of Taiwan (excluding Anthostomella). Mycotaxon 73:
343-440.

Xylaria spp. nov. (China) ... 303

Fics 7-10. Xylaria jiangsuensis (HMAS 7263, holotype). 7. Stromata on dead leaf; 8. Section of
stroma; 9-10. Asci and ascospores.

Ju YM, Hsieh HM. 2007. Xylaria species associated with nests of Odontotermes formosanus in
Taiwan. Mycologia 99: 936-957. http://dx.doi.org/10.3852/mycologia.99.6.936

Li YX, Li HJ. 1994. A novel species of Xylaria. J. Nanjing Agr. Univ. 17(3): 145-147.

Ma HX, Vasilyeva LN, Li Y. 2011a. A new species of Xylaria from China. Mycotaxon 116: 151-155.
http://dx.doi.org/10.5248/116.151

Ma HX, Vasilyeva LN, Li Y. 2011b. Xylaria choui, a new species from China. Sydowia 63: 79-83.

Ma HX, Vasilyeva LN, Li Y. 2012. The genus Xylaria in the south of China - 3. X. atroglobosa sp.
nov. Mycotaxon, 119: 381-384. http://dx.doi.org/10.5248/119.381

Ma HX, Vasilyeva LN, Li Y. 2013. The genus Xylaria in the south of China - 4. X. hemisphaerica sp.
nov. from Yunnan Province. Mycosystema 32: 602-605.

304 ... Huang & al.

Rogers JD, Ju YM. 1998. Keys to the Xylariaceae (excluding Anthostomella) of the British Isles. Bot
J Scot 50: 153-160. http://dx.doi.org/10.1080/037466098086849 12

Tai FL. 1979. Sylloge Fungorum Sinicorum. Science Press, Beijing. 1527 p.

Teng SC. 1963. Fungi of China. Science Press, Beijing. 808 p.

Whalley MA, Ju YM, Rogers JD, Whalley AJS. 2000. New xylariaceous fungi from Malaysia.
Mycotaxon 74: 135-140.

Xu AS. 1999. A new species of Xylaria. Mycosystema 18(2): 137-140

Zhu YF, Guo L. 2011. Xylaria hainanensis sp. nov. (Xylariaceae) from China. Mycosystema 30:
526-528.

MY COTAXON

http://dx.doi.org/10.5248/130.305
Volume 130, pp. 305-306 January-March 2015

NOMENCLATURAL NOVELTIES AND TYPIFICATIONS

PROPOSED IN MYCOTAXON 130-1

Ambomucor ovalisporus X.Y. Liu & R.Y. Zheng, p. 166

Ambomucor seriatoinflatus var. longior X.Y. Liu & R.Y. Zheng, p. 168
Antherospora sukhomlyniae K.G. Savchenko, p. 57

Anungitea guangxiensis J.W. Xia & X.G. Zhang, p. 43

Bertia hainanensis Lar.N. Vassiljeva, H.X. Ma, Chernyshev &
S.L. Stephenson, p. 198

Cladonia pseudofissa (Asahina) Ahti, Pino-Bodas & S. Stenroos, p. 98
Coccomyces neolitseae Qing Li & Y.R. Lin, p. 290

Coccomyces prominens Y.F. Xu &Y.R. Lin, p. 74

Datronia ustulatiligna Har. Kaur, G. Kaur & Dhingra, p. 295
Dentipellicula austroafricana Jia J. Chen, L.L. Shen & Y.C. Dai, p. 20
Ellisembia longchiensis J.W. Xia & X.G. Zhang, p. 44

Fistulina subhepatica B.K. Cui & J. Song, p. 53

Hyphoderma hallenbergii Man. Kaur, Avneet P. Singh & Dhingra, p. 223
Lactarius indochrysorrheus K. Das & Verbeken, p. 110

Lactarius olivaceoglutinus K. Das & Verbeken, p. 114

Lactarius pyriodorus K. Das & Verbeken, p. 118

Lactarius yumthangensis K. Das & Verbeken, p. 124

Leiorreuma crassimarginatum Z.F. Jia, p. 248

Lophodermium heterocladum H.L. Gu & Y.R. Lin, p. 192
Perenniporia koreana Y. Jang & J.J. Kim, p. 174

Periconiella liquidambaricola U. Braun, S. Bien & Honig, p. 254

Periconiella qualeicola Dorn.-Silva & Dianese, p. 257
= Periconiella longispora Dorn.-Silva & Dianese 2004, nom. illeg.
non P. longispora Kamal, Suj. Singh & R.P. Singh 1979

Phyllachora hainanensis Na Liu & G. Huang, p. 237
Pseudoplectania lignicola Glejdura, V. Kucera, Lizon & Kunca, p. 2
Rosellinia brunneola Wei Li bis & L. Guo, p. 233

Sarcoporia longitubulata Vlasak & Spirin, p. 282

306 ... MYCOTAXON 130-1

Terriera aequabilis Qing Li & Y.R. Lin, p. 28

Tuber xanthomonosporum Qing & Y. Wang, p. 64

Verticicladus hainanensis M.T. Guo & Z.F. Yu, p. 276

Xylaria jiangsuensis Rui S. Wang & L. Guo, p. 302

Xylaria semiglobosa G. Huang & L. Guo, p. 299

Xylaria sphaerica G. Huang & L. Guo, p. 300

Xylaria thailandica Srihanant, Petcharat & Lar.N. Vassiljeva, p. 228

Zasmidium cassines Kharwar, Arch. Singh, Raghv. Singh & Sham. Kumar, p. 242
Zasmidium fabaceicola Kharwar, Arch. Singh, Raghv. Singh & Sham. Kumar, p.244

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